Novel compositions and processes for analyte detection, quantification and amplification

ABSTRACT

This invention provides novel compositions and processes for analyte detection, quantification and amplification. Nucleic acid arrays and libraries of analytes are usefully incorporated into such compositions and processes. Universal detection elements, signaling entities and the like are employed to detect and if necessary or desirable, to quantify analytes. Amplification of target analytes are also provided by the compositions and processes of this invention.

FIELD OF THE INVENTION

[0001] This invention relates to the field of analyte detection,quantification and amplification, including compositions and processesdirected thereto.

[0002] All patents, patent applications, patent publications, scientificarticles and the like, cited or identified in this application arehereby incorporated by reference in their entirety in order to describemore fully the state of the art to which the present invention pertains.

BACKGROUND OF THE INVENTION

[0003] The quantification of RNA expression provides major insights intoanalysis of cellular metabolism, function, growth and interactions.Although individual RNA species have historically been the subject ofthese studies, more interest is currently being shown in analysis of thepatterns of the simultaneous expression of multiple RNA species of bothknown and unknown function. This approach allows comparative studies onthe patterns of expression between different populations of cells,thereby serving as an indicator of the differences in biochemicalactivities taking place within these populations. For instance, a singlegroup of cells can be divided up into two or more populations where onegroup serves as a control and the other part is exposed to drugs,metabolites or different physical conditions. In this way, although themajority of the various species of mRNA show little or no differences inexpression levels, certain mRNA species may show dramatic increased ordecreased levels of expression compared to the untreated or normalcontrol.

[0004] As an example, it has long been known that the application of aphorbol ester (PMA) results in changes in a large number ofcharacteristics of mammalian cells growing in vitro. In an experimentreported by Lockhart et al., (1996, Nature Biotechnology 14; 1675-1680)cells growing in culture were exposed to PMA and at various timesafterwards, mRNA was extracted and used to create a library of labeledprobes. This material was subsequently hybridized to an array of nucleicacids that was complementary to various mRNA sequences. Significantchanges could be seen in both the timing and the amount of induction ofvarious cellular cytokines. On the other hand, so called “house-keeping”genes such as actin and GAPDH remained essentially unaffected by thetreatment. This example demonstrates that the various mRNA's can beindependently monitored to determine which particular genes may beaffected by a treatment.

[0005] Natural differences between cell populations can also beexamined. For instance, differences in the expression levels of variousgenes can be observed when cells progress through cell cycles (Cho etal., 1 998 Mol Cell 2; 65-73 and Spellman et al., 1998 Mol. Biol. Cell95; 14863-14868). The gene expression profiles that were generated bythese studies validated this approach when significant differences inexpression were observed for genes that had previously beencharacterized as encoding cell cycle related proteins. In addition, thearrays used in these studies comprised nucleic acid sequences thatrepresented the entire genetic complement of the yeast being studied. Assuch, one of the results of these studies was the observation of anumber of genes of previously unknown function that also displayed cellcycle dependent expression. Re-examination of these particular genes byother more conventional methods demonstrated that they were involved incell cycle progression. Thus, this method was demonstrated as beingcapable of recognizing genes previously known for differentialexpression and also for identifying new genes.

[0006] The differences between normal and transformed cells have alsobeen a subject of long standing interest. The nature of the particulargenes that are either overexpressed or underexpressed relative to normalcells may provide information on the origination, progression ortreatment of cancerous cells. Array analysis has been carried out byusing RNA from tumor derived cells in comparison with expression fromnormal cells. In one study by Perou et al (1999 Proc. Nat Acad. Sci. USA96; 9212-9217) human mammary epithelial cells (HMEC) were compared withspecimens from primary breast tumors. Included in this study wereresponses to various cell factors as well as the results of confluenceor senescence in the control cultures. All of these are factors that maybe involved or affected by cellular transformation into the cancerousstate. The amount of data generated in this type of study is almostoverwhelming in its complexity. However distinct patterns or clusters ofexpression can be observed that are correlated to factors associatedwith the specimens. Further understanding will also be gained when datais gathered from expression in other tumor types and their untransformedequivalents.

[0007] There are two distinct elements in all of the expression studiesthat employ arrays. The first element is concerned with the preparationof the bank of probes that will be used to bind or capture labeledmaterial that is derived from the mRNAs that are being analyzed. Thepurpose of these arrays is to provide a multiplicity of individualprobes where each probe is located in a discrete spatially definedposition. After hybridization of the sample is carried out, theparticular amount of sample is measured for each site giving a relativemeasurement of how much material is present in the sample that hashomology with the particular probe that is located at that site. The twomost commonly used methods for array assembly operate on two verydifferent scales for synthesis of arrays.

[0008] On the simplest level of construction, discrete nucleic acids areaffixed to solid matrixes such as glass slides or nylon membranes in aprocess that is very similar to that employed by ink jet printers (Forexample, see Okamoto et al., 2000, Nature Biotechnology 18; 438-441).The nature of the probe deposited on the matrix can range from smallsynthetic oligonucleotides to large nucleic acid segments from clones.Preparation of a cloned segment to be used in this form of arrayassembly can range from E. coli colonies containing individual clonesthat are lysed and fixed directly onto a matrix or more elaborately byusing individual plasmids as templates for preparation of PCR amplifiedmaterial. The latter method is preferred due to the higher purity of thenucleic acid product. The choice of a particular probe to be used in theassembly can be directed in the sense that the function and sequence isknown. This of course will always be true when oligonucleotides are usedas the probes since they must be synthesized artificially. On the otherhand, when the probes are derived from larger cloned segments of DNA,they can be used irrespective of knowledge of sequence or function. Forinstance, a bank of probes that represent the entire yeast genome wasused in the studies cited earlier on differential expression during cellcycle progression. For human sequences, the burgeoning growth of thehuman sequencing project has provided a wealth of sequence informationthat is constantly expanding. Therefore, a popular source of probes thatcan be used to detect human transcripts has been Expressed Sequence Tags(ESTs) (Adams et al., 1991 Science 252; 1651-1656). The use of sequencesof unknown function has the advantage of a lack of any a prioriassumption concerning responsiveness in a comparative study and in fact,the study in itself may serve to identify functionality. At present,filter and glass arrays are commercially available from a number ofsources for the analysis of expression from various human tissues,developmental stages and disease conditions. On the other hand,directions for making custom arrays are widely disseminated throughoutthe literature and over the Internet.

[0009] At the other end of the scale in complexity is a process where insitu synthesis of oligonucleotides is carried out directly on a solidmatrix using a “masking” technology that is similar to that employed inetching of microcircuits (Pirrung et al., U.S. Pat. No. 5,143,854,hereby incorporated by reference). Since this process can be carried outon a very small microscale, a very large number of different probes canbe loaded onto a single “biochip” as a high density array. However,since this method depends upon site-specific synthesis, onlyoligonucleotides are used and the probes are necessarily of limitedsize. Also, since directed sequence synthesis is used, sequenceinformation has to be available for each probe. An advantage of thissystem is that instead of a single probe for a particular gene product,a number of probes from different segments can be synthesized andincorporated into the design of the array. This provides a redundancy ofinformation, establishing that changes in levels of a particulartranscript are due to fluctuations in the intended target rather than bytranscripts with one or more similar sequences. These “biochips” arecommercially available as well as the hardware and software required toread them.

[0010] Although solid supports such as plastic and glass have beencommonly used for fixation of nucleic acids, porous materials have alsobeen used. For example, oligonucleotides were joined to aldehyde groupsin polyacrylamide (Yershov et al., (1996) Proc Nat. Acad. Sci USA 93;4913-4918) and agarose (Afanassiev et al. (2000) Nucl. Acids Res. 28;e66) to synthesize arrays that were used in hybridization assays.

[0011] The second element involved in array analysis is the means bywhich the presence and amount of labeled nucleic acids bound to thevarious probes of the array will be detected. There are three levels ofuse of the target mRNA that can provide signal generation. In the firstapproach, the native RNA itself can be labeled. This has been carriedout enzymatically by phosphorylation of fragmented RNA followed by T4RNA ligase mediated addition of a biotinylated oligomer to the 5′ ends(Lockhart et al, 1996). This method has the limitation that it entailsan overnight incubation to insure adequate joining of labels to the RNA.For chemical labeling of RNA, the fragments can be labeled with psoralenthat has been linked to biotin (Lockhart et al, 1996). This method hasthe disadvantage that the crosslinking that joins the label to the RNAcan also lead to intrastrand crosslinking of target molecules reducingthe amount of hybridizable material.

[0012] In the second approach, rather than labeling the transcriptitself, the RNA is used as a template to synthesize cDNA copies by theuse of either random primers or by oligo dT primers. Extension of theprimers by reverse transcriptase can be carried out in the presence ofmodified nucleotides, thereby labeling all of the nascent cDNA copies.The modified nucleotides can have moieties attached that generatesignals in themselves or they may have moieties suitable for attachmentof other moieties capable of generation of signals. Examples of groupsthat have been used for direct signal generation have been radioactivecompounds and fluorescent compounds such as fluorescein, Texas red, Cy3and Cy 5. Direct signal generation has the advantage of simplicity buthas the limitation that in many cases there is reduced efficiency forincorporation of the labeled nucleotides by a polymerase. Examples ofgroups that have been used for indirect signal generation in arrays aredinitrophenol (DNP) or biotin ligands. Their presence is detected laterby the use of labeled molecules that have affinities for these ligands.Avidin or strepavidin specifically bind to biotin moieties andantibodies can be used that are specific for DNP or biotin. Theseproteins can be labeled themselves or serve as targets for secondarybindings with labeled compounds. Alternatively, when the labelednucleotides contain chemically active substituents such as allylaminemodifications, post-synthetic modification can be carried out by achemical addition of a suitably labeled ester.

[0013] The synthesis of a cDNA copy from an mRNA template essentiallyresults in a one to one molar ratio of labeled product compared tostarting material. In some cases there may be limiting amounts of themRNA being analyzed and for these cases, some amplification of thenucleic acid sequences in the sample may be desirable. This has led tothe use of the third approach, where the cDNA copy derived from theoriginal mRNA template is in itself used as a template for furthersynthesis. A system termed “Transcription Amplification System” (TAS)was described (Kwoh, D. Y. and Gingeras, T. R., 1989, Proc. Nat. Acad.Sci., 86, 1173-1177) in which a target specific oligonucleotide is usedto generate a cDNA copy and a second target specific oligonucleotide isused to convert the single stranded DNA into double-stranded form. Byinclusion of a T7 promoter sequence into the first oligonucleotide, thedouble-stranded molecule can be used to make multiple transcriptionproducts that are complementary to the original mRNA of interest. Thepurpose of this system was for amplification of a discrete sequence froma pool of various RNA species. No suggestion or appreciation of such asystem for the use of non-discrete primer sequences for generalamplification was described in this work.

[0014] Multiple RNA transcript copies homologous to the original RNApopulation has been disclosed by van Gelder et al. in U.S. Pat. No.5,891,636 where specific reference is given to the utility of such asystem for creating a library of various gene products in addition todiscrete sequences. Since each individual mRNA molecule has thepotential for ultimately being the source of a large number ofcomplementary transcripts, this system enjoys the advantages of linearamplification such that smaller amounts of starting material arenecessary compared to direct labeling of the original mRNA or its cDNAcopy.

[0015] However, the work described in U.S. Pat. No. 5,891,636specifically teaches away from addition of exogenous primers forsynthesis of a 2^(nd) strand. Instead, it discloses the use ofoligonucleotide primers for production of only the first strand of cDNA.For synthesis of the second strand, two possible methods were disclosed.In the first method, the nicking activity of RNase H on the originalmRNA template was used to create primers that could use the cDNA as atemplate. In the second method, DNA polymerase was added to formhairpins at the end of the first cDNA strand that could provideself-priming. The first method has a limitation that RNase H has to beadded after the completion of the cDNA synthesis reaction and a balanceof RNase H activity has to be determined to provide sufficient nickingwithout total degradation of potential RNA primers. The second methodrequires an extra step of incubation a different polymerase besides theReverse Transcriptase and also S1 nuclease has to be added to eliminatethe loop in the hairpin structure. In addition, the formation andextension by foldback is a poorly understood system that does notoperate at high efficiency where sequences and amounts of cDNA copiesmay act as random factors.

[0016] In addition to the amplification provided by the use of RNAtranscription, PCR has been included in some protocols to carry outsynthesis of a library through the use of common primer binding sites ateach end of individual sequences (Endege et al., 1999 Biotechniques 26;542-550, Ying et al., 1999 Biotechniques 27; 410-414). These methodsshare the necessity for a machine dedicated to thermal cycling.

[0017] In addition to binding analytes from a library, the nucleic acidson an array can use the analytes as templates for primer extensionreactions. For instance, determination of Single NucleotidePolymorphisms, (SNP's) has been carried out by the use of a set ofprimers at different sites on the array that exhibit sequence variationsfrom each other (Pastinen et al., 2000, Genome Research 10; 1031-1042).The ability or inability of a template to be used for primer extensionby each set of primers is an indication of the particular sequencevariations within the analytes. More complex series of reactions havealso been carried out by the use of arrays as platforms for localizedamplification as described in U.S. Pat. No. 5,641,658 and Weslin et al.,2000, Nature Biotechnology 18; 199-204. In these particular applicationsof array technology, PCR and SDA were carried out by providing a pair ofunique primers for each individual nucleic acid target at each locus ofthe array. The presence or absence of amplification at each locus of thearray served as an indicator of the presence or absence of thecorresponding target sequences in the analyte samples.

[0018] Despite the accelerated development of the synthesis and use ofDNA microarrays in recent years, the progress in the development ofarrays of proteins or other ligands has been significantly slower eventhough such arrays are an ideal format with which to study geneexpression, as well as antibody-antigen, receptor-ligand,protein-protein interactions and other applications. In previous art,protein arrays have been used for gene expression antibody screening,and enzymatic assays (Lueking et al. (1999) Anal. Biochem. 270; 103-111;de Wildt et al., (2000) Nature Biotechnology 18; 989-994, Arenkov etal., (2000) Analytical Biochemistry 278; 123-131). Protein arrays havealso been used for high throughput ELISA assays (Mendoza et al., (1999)Biotechniques 27; 778-788) and for the detection of individual proteinsin complex solutions (Haab, et al.; (2001) Genome Biology 2; 1-13).However, the use thus far has been limited because of the inherentproblems associated with proteins. DNA is extremely robust and can beimmobilized on a solid matrix, dried and rehydrated without any loss ofactivity or function. Proteins, however, are far more difficult toutilize in array formats. One of the main problems of using proteins inan array format is the difficulty of applying the protein to a solidmatrix in a form that would allow the protein to be accessible andreactive without denaturing or otherwise altering the peptide orprotein. Also, many proteins cannot be dehydrated and must be kept insolution at all times, creating further difficulties for use in arrays.

[0019] Some methods which have been used to prepare protein arraysinclude placing the proteins on a polyacrylamide gel matrix on a glassslide that has been activated by treatment with glutaraldehyde or otherreagents (Arenkov, op. cit.). Another method has been the addition ofproteins to aldehyde coated glass slides, followed by blocking of theremaining aldehyde sites with BSA after the attachment of the desiredprotein. This method, however, could not be used for small proteinsbecause the BSA obscured the protein. Peptides and small proteins havebeen placed on slides by coating the slides with BSA and then activatingthe BSA with N,N′-disuccinimidyl carbonate (Taton et al., (2000) Science2789, 1760-1763). The peptides were then printed onto the slides and theremaining activated sites were blocked with glycine, Protein arrays havealso been prepared on poly-L-Lysine coated glass slides (Haab et al.,op. cit.) and agarose coated glass slides (Afanassiev et al., (2000)Nucleic Acids Research 28, e66). “Protein Chips” are also commerciallyavailable from Ciphergen (Fremont, Calif.) for a process where proteinsare captured onto solid surfaces and analyzed by mass spectroscopy.

[0020] The use of oligonucleotides as ‘hooks’ or ‘tags’ as identifiersfor non-nucleic acid molecules has been described in the literature. Forinstance, a library of peptides has been made where each peptide isattached to a discrete nucleic acid portion and members of the libraryare tested for their ability to bind to a particular analyte. Afterisolation of the peptides that have binding affinities, identificationwas carried out by PCR to “decode” the peptide sequence (Brenner. andLerner, (1992) Proc. Nat. Acad. Sci. USA 89; 5381-5383, Needels et al.,(1993) Proc. Nat. Acad. Sci. USA 90; 10,700-10,704). Nuceleic acidsequences have also been used as tags in arrays where selectedoligonucleotide sequences were added to primers used for singlenucleotide polymorphism genotyping (Hirschhorn, et al., (2000) Proc.Natl. Acad. Sc. USA, 97; 12164-12169). However, in this case the ‘tag’is actually part of the primer design and it is used specifically forSNP detection using a single base extension assay. A patent applicationfiled by Lohse, et al., (WO 00/32823) has disclosed the use ofDNA-protein fusions for protein arrays. In this method, the protein issynthesized from RNA transcripts which are then reverse transcribed togive the DNA sequences attached to the corresponding protein. Thissystem lacks flexibility since the technology specifically relates onlyto chimeric molecules that comprise a nucleic acid and a peptide orprotein. In addition, the protein is directly derived from the RNAsequence so that the resultant DNA sequence is also dictated by theprotein sequence. Lastly, every protein that is to be used in an arrayrequires the use of an in vitro translation system made from cellextracts, a costly and inefficient system for large scale synthesis ofmultiple probes. The use of electrochemically addressed chips for usewith chimeric compositions has also been described by Bazin and Livache1999 in “Innovation and Perspectives in solid Phase Synthesis &Recombinatorial Libraries” R. Epton (Ed.) Mayflower Scientific Limited,Birmingham, UK.

SUMMARY OF THE INVENTION

[0021] This invention provides a composition of matter that comprises alibrary of analytes, the analytes being hybridized to an array ofnucleic acids, the nucleic acids being fixed or immobilized to a solidsupport, wherein the analytes comprise an inherent universal detectiontarget (UDT), and a universal detection element (UDE) attached to theUDT, wherein the UDE generates a signal indicating the presence orquantity of the analytes, or the attachment of UDE to UDT.

[0022] This invention also provides a composition of matter thatcomprises a library of analytes, such analytes being hybridized to anarray of nucleic acids, and such nucleic acids being fixed orimmobilized to a solid support, wherein the analytes comprise anon-inherent universal detection target (UDT) and a universal detectionelement (UDE) hybridized to the UDT, and wherein the UDE generates asignal directly or indirectly to detect the presence or quantity of suchanalytes.

[0023] The present invention further provides a composition of matterthat comprises a library of analytes, such analytes being hybridized toan array of nucleic acids, and such nucleic acids being fixed orimmobilized to a solid support, wherein the hybridization between theanalytes and the nucleic acids generate a domain for complex formation,and the composition further comprises a signaling entity complexed tothe domain.

[0024] The present invention yet further provides a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of the nucleic acids of interest comprise at least one inherentuniversal detection target (UDT); and (iii) universal detection elements(UDE) which generates a signal directly or indirectly; b) hybridizingthe library (ii) with the array of nucleic acids (i) to form hybrids ifthe nucleic acids of interest are present; c) contacting the UDEs withthe UDTs to form a complex bound to the array; d) detecting orquantifying the more than one nucleic acid of interest by detecting ormeasuring the amount of signal generated from UDEs bound to the array.

[0025] Also provided by this invention is a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of the nucleic acids of interest comprise at least one inherentuniversal detection target (UDT); and (iii) universal detection elements(UDE) which generates a signal directly or indirectly; b) contacting theUDEs with the UDTs in the library of nucleic acid analytes to form oneor more complexes; c) hybridizing the library of nucleic acid analyteswith the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; d) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array.

[0026] Also provided herein is a process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof a) providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of the nucleic acidsof interest comprise at least one non-inherent universal detectiontarget (UDT), wherein the non-inherent UDT is attached to the nucleicacid analytes; and (iii) universal detection elements (UDE) whichgenerate a signal directly or indirectly; b) hybridizing the library(ii) with the array of nucleic acids (i) to form hybrids if the nucleicacids of interest are present; c) contacting the UDEs with the UDTs toform a complex bound to the array; d) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array.

[0027] Another aspect provided by this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of such nucleic acids of interest comprise at least onenon-inherent universal detection target (UDT), wherein the non-inherentUDTs are attached to the nucleic acid analytes; and (iii) universaldetection elements (UDE) which generate a signal directly or indirectly;b) contacting the UDEs with the UDTs in the library of nucleic acidanalytes to form one or more complexes; c) hybridizing the library (ii)with the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; d) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array.

[0028] Another aspect provided by this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)means for attaching one or more universal detection targets (UDT) to anucleic acid; (iv) universal detection elements (UDE) which generates asignal directly or indirectly; b) attaching such UDTs (iii) to thelibrary of nucleic acid analytes (ii); c) hybridizing the library (ii)with the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; d) contacting the UDEs with the UDTs toform a complex bound to the array; e) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array.

[0029] Still another feature is process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof a) providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore universal detection targets (UDT) to a nucleic acid; (iv) universaldetection elements (UDE) which generate a signal directly or indirectly;b) attaching the UDTs (iii) to the library of nucleic acid analytes(ii); c) contacting the UDEs with the UDTs in the library of nucleicacid analytes to form one or more complexes; d) hybridizing the library(ii) with the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; e) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array.

[0030] The present invention provides additionally a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; and (iii)universal detection elements (UDEs) which bind to a domain formed bynucleic acid hybrids for complex formation and generate a signaldirectly or indirectly; b) hybridizing the library (ii) with the arrayof nucleic acids (i) to form hybrids if such nucleic acids of interestare present, wherein any formed hybrids generate a domain for complexformation; c) contacting the UDEs with any hybrids to form a complexbound to the array; d) detecting or quantifying the more than onenucleic acid of interest by detecting or measuring the amount of signalgenerated from UDEs bound to the array.

[0031] Also provided herein is a composition of matter comprising alibrary of first nucleic acid analyte copies, such first nucleic acidcopies being hybridized to an array of nucleic acids, those nucleicacids being fixed or immobilized to a solid support, wherein such firstnucleic acid copies comprise an inherent universal detection target(UDT) and a universal detection element (UDE) attached to the UDT,wherein the UDE generates a signal directly or indirectly to detect thepresence or quantity of any analytes.

[0032] Another embodiment of this invention is a composition of mattercomprising a library of first nucleic acid analyte copies, such firstnucleic acid copies being hybridized to an array of nucleic acids, thenucleic acids being fixed or immobilized to a solid support, whereinsuch first nucleic acid copies comprise one or more non-inherentuniversal detection targets (UDTs) and one or more universal detectionelements (UDEs) attached to the UDTs, wherein the UDEs generate a signaldirectly or indirectly to detect the presence or quantity of anyanalytes, and wherein the UDTs are either: (i) at the 5′ ends of thefirst nucleic acid copies and not adjacent to an oligot segment orsequence, or (ii) at the 3′ ends of the first nucleic acid copies, or(iii) both (i) and (ii).

[0033] This invention also concerns a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to the nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified, wherein each of such nucleic acids of interest comprise atleast one inherent universal detection target (UDT); (iii) universaldetection elements (UDE) which generate a signal directly or indirectly;and (iv) polymerizing means for synthesizing nucleic acid copies of thenucleic acids of analytes; b) synthesizing one or more first nucleicacid copies which are complementary to all or part of the nucleic acidanalytes and synthesizing sequences which are complementary to all orpart of the UDT to form a complementary UDT; c) hybridizing such firstnucleic acid copies with the array of nucleic acids (i) to form hybridsif such nucleic acids of interest are present; d) contacting the UDEswith the complementary UDTs of the first nucleic acid copies to form acomplex bound to the array; e) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array.

[0034] Another embodiment provided by this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to the nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified, wherein each of such nucleic acids of interest comprise atleast one inherent universal detection target (UDT); (iii) universaldetection elements (UDE) which generate a signal directly or indirectly;and (iv) polymerizing means for synthesizing nucleic acid copies of suchnucleic acid analytes; b) synthesizing one or more first nucleic acidcopies of such nucleic acid analytes; c) contacting the UDEs with theUDTs in the first nucleic acid copies to form one or more complexes; d)hybridizing such first nucleic acid copies with the array of nucleicacids (i) to form hybrids if such nucleic acids of interest are present;and e) detecting or quantifying the more than one nucleic acid ofinterest by detecting or measuring the amount of signal generated fromUDEs bound to the array.

[0035] An additional aspect of the present invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to the nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified; (iii) means for attaching one or more non-inherent universaldetection targets (UDT) to a nucleic acid; (iv) universal detectionelements (UDE) which generate a signal directly or indirectly; and (v)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes; b) attaching the non-inherent UDTs to either the 3′ endsof the nucleic acid analytes, the 5′ ends of the first nucleic acidanalytes, or both the 3′ ends and the 5′ ends of the nucleic acidanalytes; c) synthesizing one or more first nucleic acid copies of thenucleic acid analytes; d) hybridizing the first nucleic acid copies withthe array of nucleic acids (i) to form hybrids if such nucleic acids ofinterest are present; e) contacting the UDEs with the UDTs of the firstnucleic acid copies to form a complex bound to the array; f) detectingor quantifying the more than one nucleic acid of interest by detectingor measuring the amount of signal generated from UDEs bound to thearray.

[0036] Also provided herein is a process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof a) providing (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to the nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) means for attachingone or more non-inherent universal detection targets (UDT) to a nucleicacid; (iv) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (v) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes; b) attaching suchnon-inherent UDTs to either the 3′ ends of the nucleic acid analytes,the 5′ ends of the first nucleic acid analytes, or both the 3′ ends andthe 5′ ends of the nucleic acid analytes; c) synthesizing one or morefirst nucleic acid copies of the nucleic acid analytes; d) contactingthe UDEs with the UDTs of the first nucleic acid copies to formcomplexes; e) hybridizing the first nucleic acid copies with the arrayof nucleic acids (i) to form hybrids if any nucleic acids of interestare present; f) detecting or quantifying the more than one nucleic acidof interest by detecting or measuring the amount of signal generatedfrom UDEs bound to the array.

[0037] Another embodiment provided herein is a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to such nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified; (iii) means for attaching one or more non-inherent universaldetection targets (UDT) to a nucleic acid; (iv) universal detectionelements (UDE) which generate a signal directly or indirectly; and (v)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes; b) synthesizing one or more first nucleic acid copies ofthe nucleic acid analytes; c) attaching the non-inherent UDTs to eitherthe 3′ ends of the first nucleic acid copies, the 5′ ends of the firstnucleic acid copies, or both the 3′ ends and the 5′ ends of the firstnucleic acid copies; d) hybridizing the first nucleic acid copies withthe array of nucleic acids (i) to form hybrids if any nucleic acids ofinterest are present; e) contacting the UDEs with the UDTs of the firstnucleic acid copies to form a complex bound to the array; and f)detecting or quantifying the more than one nucleic acid of interest bydetecting or measuring the amount of signal generated from UDEs bound tothe array.

[0038] Another process provided by this invention is for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to the nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified; (iii) means for attaching one or more non-inherent universaldetection targets (UDT) to a nucleic acid; (iv) universal detectionelements (UDE) which generate a signal directly or indirectly; and (v)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes; b) synthesizing one or more first nucleic acid copies ofthe nucleic acid analytes; c) attaching the non-inherent UDTs to eitherthe 3′ ends of the first nucleic acid copies, the 5′ ends of the firstnucleic acid copies, or both the 3′ ends and the 5′ ends of the firstnucleic acid copies; d) contacting the UDEs with the UDTs of the firstnucleic acid copies to form a complex; e) hybridizing the first nucleicacid copies with the array of nucleic acids (i) to form hybrids if anynucleic acids of interest are present; and f) detecting or quantifyingthe more than one nucleic acid of interest by detecting or measuring theamount of signal generated from UDEs bound to the array.

[0039] Yet further provided is a process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof a) providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) universal detection elements(UDEs) which bind to a domain for complex formation formed by nucleicacid hybrids and generate a signal directly or indirectly; and (iv)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes; b) synthesizing one or more nucleic acid copies of thenucleic acid analytes; c) hybridizing the first nucleic acid copies withthe array of nucleic acids (i) to form hybrids if any nucleic acids ofinterest are present, wherein any formed hybrids generate a domain forcomplex formation; d) contacting the UDEs with the hybrids to form acomplex bound to the array; and e) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array.

[0040] Another aspect provided by this invention is a composition ofmatter comprising a library of double-stranded nucleic acidssubstantially incapable of in vivo replication and free of non-inherenthomopolymeric sequences, the nucleic acids comprising sequencescomplementary or identical in part or whole to inherent sequences of alibrary obtained from a sample, wherein the double-stranded nucleicacids comprise at least one inherent universal detection target (UDT)proximate to one end of the double strand and at least one non-inherentproduction center proximate to the other end of the double strand.

[0041] Yet another aspect of this invention concerns a composition ofmatter comprising a library of double-stranded nucleic acidssubstantially incapable of in vivo replication, such nucleic acidscomprising sequences complementary or identical in part or whole toinherent sequences of a library obtained from a sample, wherein thedouble-stranded nucleic acids comprise at least four (4) non-inherentnucleotides proximate to one end of the double strand and a non-inherentproduction center proximate to the other end of the double strand.

[0042] Among other useful aspects of this invention is a composition ofmatter comprising a library of double-stranded nucleic acids fixed to asolid support, those nucleic acids comprising sequences complementary oridentical in part or whole to inherent sequences of a library obtainedfrom a sample and the nucleic acids further comprising at least onefirst sequence segment of non-inherent nucleotides proximate to one endof the double strand and at least one second sequence segment proximateto the other end of the double strand, the second sequence segmentcomprising at least one production center.

[0043] Another feature of this invention is a composition of mattercomprising a library of double-stranded nucleic acids attached to asolid support, the nucleic acids comprising sequences complementary oridentical in part or whole to inherent sequences of a library obtainedfrom a sample, wherein the double-stranded nucleic acids comprise atleast one inherent universal detection target (UDT) proximate to one endof the double strand and at least one non-inherent production centerproximate to the other end of the double strand.

[0044] The invention herein also provides a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, the polymerizing meanscomprising a first set of primers and a second set of primers, whereinthe second set of primers comprises at least two segments, the firstsegment at the 3′ end comprising random sequences, and the secondsegment comprising at least one production center; (iv) means forsynthesizing nucleic acid copies under isothermal or isostaticconditions; b) contacting the library of nucleic acid analytes with thefirst set of primers to form more than one first bound entity; c)extending the bound first set of primers by means of template sequencesprovided by the nucleic acid analytes to form first copies of theanalytes; d) contacting the extended first copies with the second set ofprimers to form more than one second bound entity; e) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; f)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; g) hybridizing any nucleic acid copies formed instep f) to the array of nucleic acids provided in step a) (i); and h)detecting or quantifying any of the hybridized copies obtained in stepg).

[0045] Also provided by this invention is a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set of primers comprise at least one productioncenter; and (iv) means for synthesizing nucleic acid copies underisothermal or isostatic conditions; b) contacting the library of nucleicacid analytes with the first set of primers to form more than one firstbound entity; c) extending the bound first set of primers by means oftemplate sequences provided by the nucleic acid analytes to form firstcopies of the analytes; d) extending the first copies by means of atleast four (4) or more non-inherent homopolymeric nucleotides; e)contacting the extended first copies with the second set of primers toform more than one second bound entity; f) extending the bound secondset of primers by means of template sequences provided by the extendedfirst copies to form more than one complex comprising extended firstcopies and extended second set of primers; g) synthesizing from aproduction center in the second set of primers in the complexes one ormore nucleic acid copies under isothermal or isostatic conditions; h)hybridizing the nucleic acid copies formed in step g) to the array ofnucleic acids provided in step a) (i); and i) detecting or quantifyingany of the hybridized copies obtained in step h).

[0046] Another feature of this invention is a process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set comprises at least one production center; (iv) aset of oligonucleotides or polynucleotides complementary to at least onesegment or sequence of the second set of primers; and(v) means forligating the set of oligonucleotides or polynucleotides (iv); b)contacting the library of nucleic acid analytes with the first set ofprimers to form more than one first bound entity; c) extending the boundfirst set of primers by means of template sequences provided by thenucleic acid analytes to form first copies of the analytes; d) ligatingthe set of oligonucleotides or polynucleotides a) (iv) to the 3′ end ofthe first copies formed in step c) to form more than one ligatedproduct; e) contacting the ligated product with the second set ofprimers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe ligated products formed in step d) to form more than one complexcomprising the ligated products and the extended second set of primers;g) synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph).

[0047] Still yet further this invention provides a process for detectingor quantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the second set comprises at least one production center; (iv) aset of oligonucleotides or polynucleotides complementary to at least onesegment or sequence of the second set of primers; and (v) means forligating the set of oligonucleotides or polynucleotides (iv); b)contacting the library of nucleic acid analytes with the first set ofprimers to form more than one first bound entity; c) extending the boundfirst set of primers by means of template sequences provided by thenucleic acid analytes to form first copies of the analytes; d) ligatingthe set of oligonucleotides or polynucleotides a) (iv) to the 3′ end ofthe first copies formed in step c) to form more than one ligatedproduct; e) contacting the ligated product with the second set ofprimers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe ligated products formed in step d) to form more than one complexcomprising the ligated products and the extended second set of primers;g) synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph).

[0048] Still yet further provided by this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers, a second set of primers and athird set of primers wherein the third set comprises at least oneproduction center; and b) contacting the library of nucleic acidanalytes with the first set of primers to form a first set of boundprimers; c) extending the first set of bound primers by means oftemplate sequences provided by the nucleic acid analytes to form firstcopies of the analytes; d) contacting the extended first copies with thesecond set of primers to form a second set of bound primers; e)extending the second set of bound primers by means of template sequencesprovided by the extended first copies to form second copies of thenucleic acid analytes; f) contacting the second copies with the thirdset of primers to form more than one third bound entity to form a thirdset of bound primers; g) extending the third set of bound primers bymeans of template sequences provided by the extended second set ofprimers to form a hybrid comprising a second copy, a third copy and atleast one production center; h) synthesizing from the production centerin the second set of primers in the complexes one or more nucleic acidcopies under isothermal or isostatic conditions; i) hybridizing thenucleic acid copies formed in step i) to the array of nucleic acidsprovided in step a) (i); and j) detecting or quantifying any of thehybridized copies obtained in step i).

[0049] Also uniquely provided in this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set of primers are fixed or immobilized to a solidsupport, and wherein the second set of primers comprises at least twosegments, the first segment at the 3′ end comprising random sequences,and the second segment comprising at least one production center; (iv)means for synthesizing nucleic acid copies under isothermal or isostaticconditions; b) contacting the library of nucleic acid analytes with thefirst set of primers to form more than one first bound entity; c)extending the bound first set of primers by means of template sequencesprovided by the nucleic acid analytes to form first copies of theanalytes; d) contacting the extended first copies with the second set ofprimers to form more than one second bound entity; e) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; f)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; g) hybridizing the nucleic acid copies formed instep f) to the array of nucleic acids provided in step a) (i); and h)detecting or quantifying any of the hybridized copies obtained in stepg).

[0050] Another significant aspect of this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set of primers are fixed or immobilized to a solidsupport, and wherein the first set of primers comprise at least oneproduction center; and (iv) means for synthesizing nucleic acid copiesunder isothermal or isostatic conditions; b) contacting the library ofnucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) extending the first copies bymeans of at least four (4) or more non-inherent homopolymericnucleotides; e) contacting the extended first copies with the second setof primers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; g)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph).

[0051] Also provided in accordance with the present invention is aprocess for detecting or quantifying more than one nucleic acid ofinterest in a library comprising the steps of a) providing (i) an arrayof fixed or immobilized nucleic acids identical or complementary in partor whole to sequences of the nucleic acids of interest; (ii) a libraryof nucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) polymerizing means forsynthesizing nucleic acid copies of the nucleic acid analytes, suchpolymerizing means comprising a first set of primers and a second set ofprimers, wherein the first set of primers are fixed or immobilized to asolid support, and wherein the first set comprises at least oneproduction center; (iv) a set of oligonucleotides or polynucleotidescomplementary to at least one segment or sequence of the second set ofprimers; and (v) means for ligating the set of oligonucleotides orpolynucleotides (iv); b) contacting the library of nucleic acid analyteswith the first set of primers to form more than one first bound entity;c) extending the bound first set of primers by means of templatesequences provided by the nucleic acid analytes to form first copies ofthe analytes; d) ligating the set of oligonucleotides or polynucleotidesa) (iv) to the 3′ end of the first copies formed in step c) to form morethan one ligated product; e) contacting the ligated product with thesecond set of primers to form more than one second bound entity; f)extending the bound second set of primers by means of template sequencesprovided by the ligated products formed in step d) to form more than onecomplex comprising the ligated products and the extended second set ofprimers; g) synthesizing from a production center in the second set ofprimers in the complexes one or more nucleic acid copies underisothermal or isostatic conditions; h) hybridizing the nucleic acidcopies formed in step g) to the array of nucleic acids provided in stepa) (i); and i) detecting or quantifying any of the hybridized copiesobtained in step h).

[0052] Another feature of the present invention concerns a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set of primers are fixed or immobilized to a solidsupport, and wherein the second set i;:;; comprises at least oneproduction center; (iv) a set of oligonucleotides or polynucleotidescomplementary to at least one segment or sequence of the second set ofprimers; and (v) means for ligating the set of oligonucleotides orpolynucleotides (iv); b) contacting the library of nucleic acid analyteswith the first set of primers to form more than one first bound entity;c) extending the bound first set of primers by means of templatesequences provided by the nucleic acid analytes to form first copies ofthe analytes; d) ligating the set of oligonucleotides or polynucleotidesa) (iv) to the 3′ end of the first copies formed in step c) to form morethan one ligated product; e) contacting the ligated product with thesecond set of primers to form more than one second bound entity; f)extending the bound second set of primers by means of template sequencesprovided by the ligated products formed in step d) to form more than onecomplex comprising the ligated products and the extended second set ofprimers; g) synthesizing from a production center in the second set ofprimers in the complexes one or more nucleic acid copies underisothermal or isostatic conditions; h) hybridizing the nucleic acidcopies formed in step g) to the array of nucleic acids provided in stepa) (i); and i) detecting or quantifying any of the hybridized copiesobtained instep h).

[0053] Yet another process is provided by this invention, the processbeing one for detecting or quantifying more than one nucleic acid ofinterest in a library and comprising the steps of a) providing (i) anarray of fixed or immobilized nucleic acids identical or complementaryin part or whole to sequences of the nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; and (iii) polymerizingmeans for synthesizing nucleic acid copies of the nucleic acid analytes,such polymerizing means comprising a first set of primers, a second setof primers and a third set of primers, wherein the first set of primersare fixed or immobilized to a solid support, and wherein the third setcomprises at least one production center; and b) contacting the libraryof nucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) contacting the extended firstcopies with the second set of primers to form more than one second boundentity; e) extending the bound I second set of primers by means oftemplate sequences provided by the extended first copies to form anextended second set of primers; f) separating the extended second set ofprimers obtained in step e); g) contacting the extended second set ofprimers with the third set of primers to form more than one third boundentity; h) extending the third bound entity by means of templatesequences provided by the extended second set of primers to form morethan one complex comprising the extended third bound entity and theextended set of primers; i) synthesizing from a production center in thesecond set of primers in the complexes one or more nucleic acid copiesunder isothermal or isostatic conditions; j) hybridizing the nucleicacid copies formed in step i) to the array of nucleic acids provided instep a) (i); and k) detecting or quantifying any of the hybridizedcopies obtained in step j).

[0054] Another significant embodiment provided herein is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to sequences of thenucleic acids of interest; (ii) a library of nucleic acid analytes whichmay contain the nucleic acids of interest sought to be detected orquantified; and (iii) polymerizing means for synthesizing nucleic acidcopies of the nucleic acid analytes, such polymerizing means comprisinga first set of primers; b) contacting the nucleic acid analytes with thefirst set of primers to form a first bound entity; c) extending thebound set of first set of primers by means of template sequencesprovided by the nucleic acid analytes to form first nucleic acid copiesof the analytes; d) separating the first nucleic acid copies from theanalytes; e) repeating steps b), c) and d) until a desirable amount offirst nucleic acid copies have been synthesized; f) hybridizing thenucleic nucleic acid copies formed in step e) to the array of nucleicacids provided in step (i); and g) detecting or quantifying any of thehybridized first nucleic acid copies obtained in step f).

[0055] The invention described herein also provides a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to sequences of thenucleic acids of interest; (ii) a library of nucleic acid analytes whichmay contain the nucleic acids of interest sought to be detected orquantified; (iii) polymerizing means for synthesizing nucleic acidcopies of the nucleic acid analytes, such polymerizing means comprisinga first set of primers and a second set of primers; (iv) means foraddition of sequences to the 3′ end of nucleic acids; b) contacting thenucleic acid analytes with the first set of primer to form a first boundentity; c) extending the bound set of first set of primers by means oftemplate sequences provided by the nucleic acid analytes to form firstnucleic acid copies of the analytes; d) extending the first nucleiccopies by the addition of non-template derived sequences to the 3′ endof the first nucleic acid copies; e) contacting the extended firstnucleic acid copies with the second set of primers to form a secondbound entity; f) extending the bound set of second set of primers bymeans of template sequences provided by the extended first nucleic acidcopies to form second nucleic acid copies; g) separating the secondnucleic acid copies from the extended first nucleic acid copies; h)repeating steps e), f) and g) until a desirable amount of second nucleicacid copies have been synthesized; i) hybridizing the second nucleicacid copies formed in step h) to the array of nucleic acids provided instep (i); and j) detecting or quantifying any of the hybridized secondnucleic acid copies obtained in step i).

[0056] Among other significant compositions provided by the presentinvention is a composition of matter that comprises an array of solidsurfaces comprising discrete areas, wherein at least two of the discreteareas each comprises a first set of nucleic acid primers; and a secondset of nucleic acid primers; wherein the nucleotide sequences in thefirst set of nucleic acid primers are different from the nucleotidesequences in the second set of nucleic acid primers; wherein thenucleotide sequences of a first set of nucleic acid primers of a firstdiscrete area and the nucleotide sequences of a first set of nucleicacid primers of a second -discrete area differ from each other by atleast one base; and wherein the nucleotide sequences of the second setof nucleic acid primers of a first discrete area and the nucleotidesequences of the second set of nucleic acid primers of a second discretearea are substantially the same or identical.

[0057] A related composition of this invention concerns a composition ofmatter that comprises an array of solid surfaces comprising a pluralityof discrete areas; wherein at least two of the discrete areas eachcomprises a first set of nucleic acid primers; and a second set ofnucleic acid primers; wherein the nucleotide sequences in the first setof nucleic acid primers are different from the nucleotide sequences inthe second set of nucleic acid primers; wherein the nucleotide sequencesof a first set of nucleic acid primers of a first discrete area and thenucleotide sequences of a first set of nucleic acid primers of a seconddiscrete area differ substantially from each other; and wherein thenucleotide sequences of the second set of nucleic acid primers of afirst discrete area and the nucleotide sequences of the second set ofnucleic acid primers of a second discrete area are substantially thesame or identical.

[0058] Related to the last-mentioned compositions are processes forproducing two or more copies of nucleic acids of interest in a librarycomprising the steps of a) providing (i) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of thediscrete areas each comprises: (1) a first set of nucleic acid primers;and (2) a second set of nucleic acid primers; wherein the nucleotidesequences in the first set of nucleic acid primers are different fromthe nucleotide sequences in the second set of nucleic acid primers;wherein the nucleotide sequences of a first set of nucleic acid primersof a first discrete area and the nucleotide sequences of a first set ofnucleic acid primers of a second discrete area differ from each other byat least one base; and wherein the nucleotide sequences of the secondset of nucleic acid primers of a first discrete area and the nucleotidesequences of the second set of nucleic acid primers of a second discretearea are substantially the same or identical; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacids of interest; b) contacting a primer of the first set with acomplementary sequence in the nucleic acid of interest; c) extending theprimer in the first set using the nucleic acid of interest as a templateto generate an extended first primer; d) contacting a primer in thesecond set with a complementary sequence in the extended first primer;e) extending the primer in the second set using the extended firstprimer as a template to generate an extended second primer; f)contacting a primer in the first set with a complementary sequence inthe extended second primer; g) extending the primer in the first setusing the extended second primer as a template to generate an extendedfirst primer; and h) repeating steps d) through g) above one or moretimes.

[0059] Another related process of the present invention is useful fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of solidsurfaces comprising a plurality of discrete areas; wherein at least twoof such discrete areas each comprises: (1) a first set of nucleic acidprimers; and (2) a second set of nucleic acid primers; wherein thenucleotide sequences in the first set of nucleic acid primers aredifferent from the nucleotide sequences in the second set of nucleicacid primers; wherein the nucleotide sequences of a first set of nucleicacid primers of a first discrete area and the nucleotide sequences of afirst set of nucleic acid primers of a second discrete area differ fromeach other by at least one base; and wherein the nucleotide sequences ofthe second set of nucleic acid primers of a first discrete area and thenucleotide sequences of the second set of nucleic acid primers of asecond discrete area are substantially the same or identical; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest; (iii) polymerizing means for synthesizing nucleic acid copiesof the nucleic acids of interest; and (iv) non-radioactive signalgenerating means capable of being attached to or incorporated intonucleic acids; b) contacting a primer of the first set with acomplementary sequence in the nucleic acid of interest; c) extending theprimer in the first set using the nucleic acid of interest as a templateto generate an extended first primer; d) contacting a primer in thesecond set with a complementary sequence in the extended first primer;e) extending the primer in the second set using the extended firstprimer as a template to generate an extended second primer; f)contacting a primer in the first set with a complementary sequence inthe extended second primer; g) extending the primer in the first setusing the extended second primer as a template to generate an extendedfirst primer; h) repeating steps d) through g) above one or more times;and i) detecting or quantifying by means of the non-radioactive signalgenerating means attached to or incorporated into any of the extendedprimers in steps c), e), g), and h).

[0060] Another useful composition provided by the present invention is acomposition of matter that comprises an array of solid surfacescomprising a plurality of discrete areas, wherein at least two of suchdiscrete areas comprise: a chimeric composition comprising a nucleicacid portion; and a non-nucleic acid portion, wherein the nucleic acidportion of a first discrete area has the same sequence as the nucleicacid portion of a second discrete area, and wherein the non-nucleic acidportion has a binding affinity for analytes of interest.

[0061] Further provided by the present invention is a composition ofmatter that comprises an array of solid surfaces comprising a pluralityof discrete areas; wherein at least two of the discrete areas comprise achimeric composition hybridized to complementary sequences of nucleicacids fixed or immobilized to the discrete areas, wherein the chimericcomposition comprises a nucleic acid portion, and a non-nucleic acidportion, the nucleic acid portion comprising at least one sequence,wherein the non-nucleic acid portion has a binding affinity for analytesof interest, and wherein when the non-nucleic acid portion is a peptideor protein, the nucleic acid portion does not comprises sequences whichare either identical or complementary to sequences that code for suchpeptide or protein.

[0062] Also provided as a significant aspect of the present invention isa process for detecting or quantifying analytes of interest, the processcomprising the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas, wherein at least two of suchdiscrete areas comprise a chimeric composition comprising a nucleic acidportion, and a non-nucleic acid portion; wherein the nucleic acidportion of a first discrete area has the same sequence as the nucleicacid portion of a second discrete area; and wherein the non-nucleic acidportion has a binding affinity for analytes of interest; b) a samplecontaining or suspected of containing one or more of the analytes ofinterest; and c) signal generating means; 2) contacting the array a)with the sample b) under conditions permissive of binding the analytesto the non-nucleic acid portion; 3) contacting the bound analytes withthe signal generating means; and 4) detecting or quantifying thepresence of the analytes.

[0063] Another feature provided by the present invention is a processfor detecting or quantifying analytes of interest, this processcomprising the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of suchdiscrete areas comprise a chimeric composition comprising a nucleic acidportion; and a non-nucleic acid portion; wherein the nucleic acidportion of a first discrete area has the same sequence as the nucleicacid portion of a second discrete area; and wherein the non-nucleic acidportion has a binding affinity for analytes of interest; b) a samplecontaining or suspected of containing one or more of the analytes ofinterest; and c) signal generating means; 2) labeling the analytes ofinterest with the signal generating means; 3) contacting the array a)with the labeled analytes under conditions permissive of binding thelabeled analytes to the non-nucleic acid portion; and 4) detecting orquantifying the presence of the analytes.

[0064] Also provided by the present invention is a process for detectingor quantifying analytes of interest, the process comprising the stepsof 1) providing a) an array of solid surfaces comprising a plurality ofdiscrete areas; wherein at least two of such discrete areas comprisenucleic acids fixed or immobilized to such discrete areas, b) chimericcompositions comprising: i) a nucleic acid portion; and ii) anon-nucleic acid portion; the nucleic acid portion comprising at leastone sequence, wherein the non-nucleic acid portion has a bindingaffinity for analytes of interest, and wherein when the non-nucleic acidportion is a peptide or protein, the nucleic acid portion does notcomprise sequences which are either identical or complementary tosequences that code for the peptide or protein; c) a sample containingor suspected of containing the analytes of interest; and d) signalgenerating means; 2) contacting the array with the chimeric compositionsto hybridize the nucleic acid portions of the chimeric compositions tocomplementary nucleic acids fixed or immobilized to the array; 3)contacting the array a) with the sample b) under conditions permissiveof binding the analytes to the non-nucleic acid portion; 4) contactingthe bound analytes with the signal generating means; and 5) detecting orquantifying the presence of the analytes.

[0065] Additionally this invention provides a process for detecting orquantifying analytes of interest, the process comprising the steps of 1)providing a) an array of solid surfaces comprising a plurality ofdiscrete areas; wherein at least two of the discrete areas comprisenucleic acids fixed or immobilized to the discrete areas, b) chimericcompositions comprising i) a nucleic acid portion; and ii) a non-nucleicacid portion, the nucleic acid portion comprising at least one sequence,wherein the non-nucleic acid portion has a binding affinity for analytesof interest, and wherein when the non-nucleic acid portion is a peptideor protein, the nucleic acid portion does not comprise sequences whichare either identical or complementary to sequences that code for thepeptide or protein; c) a sample containing or suspected of containingthe analytes of interest; and d) signal generating means; 2) contactingthe chimeric compositions with the sample b) under conditions permissiveof binding the analytes to the non-nucleic acid portion; 3) contactingthe array with the chimeric compositions to hybridize the nucleic acidportions of the chimeric compositions to complementary nucleic acidsfixed or immobilized to the array; 4) contacting the bound analytes withthe signal generating means; and 5) detecting or quantifying thepresence of the analytes.

[0066] Another useful provision of the invention herein is a process fordetecting or quantifying analytes of interest, such process comprisingthe steps of 1) providing a) an array of solid surfaces comprising aplurality of discrete areas; wherein at least two of the discrete areascomprise nucleic acids fixed or immobilized to the discrete areas, b)chimeric compositions comprising i) a nucleic acid portion; and ii) anon-nucleic acid portion; the nucleic acid portion comprising at leastone sequence, wherein the non-nucleic acid portion has a bindingaffinity for analytes of interest, and wherein when the non-nucleic acidportion is a peptide or protein, the nucleic acid portion does notcomprise sequences which are either identical or complementary tosequences that code for the peptide or protein; c) a sample containingor suspected of containing the analytes of interest; and d) signalgenerating means; 2) contacting the array with the chimeric compositionsto hybridize the nucleic acid portions of the chimeric compositions tocomplementary nucleic acids fixed or immobilized to the array; 3)labeling the analytes of interest with the signal generating means; 4)contacting the array with the labeled analytes to bind the analytes tothe non-nucleic acid portion; and 5) detecting or quantifying thepresence of the analytes.

[0067] Yet further provided by the present invention is a process fordetecting or quantifying analytes of interest, the process comprisingthe steps of 1) providing a) an array of solid surfaces comprising aplurality of discrete areas; wherein at least two of the discrete areascomprise nucleic acids fixed or immobilized to the discrete areas, b)chimeric compositions comprising: i) a nucleic acid portion; and ii) anon-nucleic acid portion; the nucleic acid portion comprising at leastone sequence, wherein the non-nucleic acid portion has a bindingaffinity for analytes of interest, and wherein when the non-nucleic acidportion is a peptide or protein, such nucleic acid portion does notcomprise sequences which are either identical or complementary tosequences that code for the peptide or protein; c) a sample containingor suspected of containing the analytes of interest; and d) signalgenerating means; 2) contacting the array with the chimeric compositionsto hybridize the nucleic acid portions of the chimeric compositions tocomplementary nucleic acids fixed or immobilized to the array; 3)labeling the analytes of interest with the signal generating means; 4)contacting the array with the labeled analytes to bind the analytes tothe non-nucleic acid portion; and 5) detecting or quantifying thepresence of the analytes.

BRIEF DESCRIPTION OF THE FIGURES

[0068]FIG. 1 shows an array with mRNA from a library of analytes withUDTs.

[0069]FIG. 2 shows fragmentation of analytes followed by addition ofnon-inherent UDTs to analytes.

[0070]FIG. 3 depicts the incorporation of a non-inherent UDT to a 1stcNA copy by means of a primer.

[0071]FIG. 4 illustrates the use of Random Primers with ProductionCenters for 2^(nd) strand synthesis.

[0072]FIG. 5 relates to the same process as FIG. 4 wherein theProduction Centers are double-stranded.

[0073]FIG. 6 illustrates 2nd cNA strand priming at terminal and internalsites.

[0074]FIG. 7 illustrates 2nd cNA strand priming after Terminaltransferase addition of homopolymeric sequences.

[0075]FIG. 8 shows the addition of primer binding sites by ligation.

[0076]FIG. 9 illustrates multiple additions of primer binding sites.

[0077]FIG. 10 shows 1 st strand synthesis by extension of an oligo dTprimer bound to a bead followed by 2nd cNA strand synthesis with randomprimers having production centers.

[0078]FIG. 11 illustrates 1st strand synthesis from poly T primerindirectly bound to a bead followed by 2nd strand synthesis with randomprimers having production center.

[0079]FIG. 12 shows the incorporation of a promoter during 3rd strandsynthesis.

[0080]FIG. 13 illustrates the synthesis of an amplicon for isothermalamplification of a library of analytes.

[0081]FIG. 14 shows the synthesis of an amplicon for SDA amplification.

[0082]FIG. 15 shows the ligation of a primer binding site for isothermalamplification.

[0083]FIG. 16 shows the binding of an analyte to an array with SPEs andUPEs for solid phase amplification.

[0084]FIG. 17 shows the extension of an SPE on an array during solidphase amplification.

[0085]FIG. 18 shows the binding of an UPE to an extended SPE followed byextension of the UPE during solid phase amplification.

[0086]FIG. 19 shows solid phase amplification in which binding ofextended SPEs and UPEs to unextended SPEs and UPEs occur.

[0087]FIG. 20 depicts an amplification array for comparative analysis.

[0088]FIG. 21 illustrates the use of an array with SPEs and UPEs for SNPanalysis.

[0089]FIG. 22 relates to binding of analytes to SPEs on an array.

[0090]FIG. 23 shows the binding of primers to extended SPEs on an array.

[0091]FIG. 24 demonstrates the binding of primers and extended primersto SPEs on an array.

[0092]FIG. 25 shows the extension of primers and SPEs on an array inaccordance with amplification disclosed in this invention.

[0093]FIG. 26 depicts the binding of nucleic acid portions of chimericcompositions to complementary sequences on an array

[0094]FIG. 27 is a gel analysis illustrating the dependency on ReverseTranscriptase for the amplification of a library in accordance with thisinvention and Example 3 below.

[0095]FIG. 28 is a gel analysis that demonstrates transcription aftermultiple rounds of 2nd strand synthesis as described further below inExample 4.

[0096]FIG. 29 is also a gel analysis that shows second round of RNAtranscription from a library as described in Example 5 below.

[0097]FIG. 30 is a gel analysis also shows transcription from librarymade after poly dG tailing in accordance with the present invention andExample 6 below.

[0098]FIG. 31 is a gel analysis that shows RNA transcription after aseries of reactions one of which was 2nd strand synthesis bythermostable DNA polymerases as described in Example 9 below.

[0099]FIG. 32 is a gel analysis that shows transcription from librariesmade from sequential synthesis of 2nd strands as further described inExample 10 below.

[0100]FIG. 33 is also a gel analysis of amplification of a library ofanalytes using various reverse transcriptases for 1st stand synthesis.

DETAILED DESCRIPTION OF THE INVENTION

[0101] The present invention discloses novel methods, compositions andkits that can be used in making and analyzing a library of nucleicacids. The nucleic acids in the sample being tested can be used directlyfor signal generation or they can be used as templates to provide one ormore nucleic acid copies that comprise sequences that are eitheridentical or complementary to the original sequences.

[0102] In the present invention the following terms are used and definedbelow:

[0103] An analyte is a biological polymer or ligand that is isolated orderived from biological sources such as organs, tissues or cells, ornon-biological sources by synthetic or enzymatic means or processes.Examples of biological polymers can include but are not limited tooligonucleotides, polynucleotides, oligopeptides, polypeptides,oligosaccharides, polysaccharides and lipids. Examples of ligands caninclude but are not necessarily limited to non-peptide antigens,hormones, enzyme substrates, vitamins, drugs, and non-peptide signalmolecules.

[0104] A library is a diverse collection of nucleic acids thatcomprises: a) analytes; b) nucleic acids derived from analytes thatcomprise sequences that are complementary to sequences in the analytes;c) nucleic acids derived from analytes that comprise sequences that areidentical to sequences in the analytes; and d) any combination of theforegoing.

[0105] A label is any moiety that is capable of directly or indirectlygenerating a signal.

[0106] A production center is a segment of a nucleic acid or analoguethereof that is capable of producing more than one copy of a sequencethat is identical or complementary to sequences that are operably linkedto the production center.

[0107] Universal Detection Targets (UDTs) are defined as common orconserved segments in diverse nucleic acids that are present inpopulations of nucleic acids in a sample and are capable of recognitionby a corresponding binding partner. The UDTs may be intrinsic or theymay be artificially incorporated into nucleic acids. Examples ofinherent UDTs can comprise but not be limited to 3′ poly A segments, 5′caps, secondary structures and consensus sequences. Examples of inherentconsensus sequences that might find use in the present invention cancomprise but not be limited to signal sites for poly A addition,splicing elements and multicopy repeats such as Alu sequences. UDTs mayalso be artificially incorporated into nucleic acids by an addition tothe original analyte nucleic acid or during synthesis of nucleic acidsthat comprise sequences that are identical or complementary to thesequences of the original analytes. Artificially added UDTs may belabeled themselves or they may serve as binding partners.

[0108] Universal Detection Elements (UDEs) are comprised of twosegments: a first segment that is capable of acting as a binding partnerfor a UDT and a second segment that is either labeled or otherwisecapable of generating a detectable signal. In some cases the first andsecond segments can be overlapping or even comprise the same segments.When UDEs are labeled, they may comprise a single signal moiety or theymay comprise more than one signal entity. Segments of UDEs involved inbinding to UDTs or signal generation may comprise but not be limited topolymeric substances such as nucleic acids, nucleic acid analogues,polypeptides, polysacharides or synthetic polymers.

[0109] The present invention discloses the use of UDTs and UDEs for thepurpose of array analysis. The present invention also discloses novelmethods for incorporation of production centers into nucleic acidlibraries that may be used in array analysis. These production centersmay provide amplification of sequences that are identical orcomplementary to sequences in the original diverse nucleic acidanalytes. The products derived from these production centers may belabeled themselves or UDTs may be incorporated for detection purposes.Nucleic acids that may be of use in the present invention can compriseor be derived from DNA or RNA. The original population of nucleic acidsmay comprise but not be limited to genomic DNA, unspliced RNA, mRNA,rRNA and snRNA.

[0110] This invention provides a composition of matter that comprises alibrary of analytes, the analytes being hybridized to an array ofnucleic acids, the nucleic acids being fixed or immobilized to a solidsupport, wherein the analytes comprise an inherent universal detectiontarget (UDT), and a universal detection element (UDE) attached to theUDT, wherein the UDE generates a signal indicating the presence orquantity of the analytes, or the attachment of UDE to UDT. The libraryof analytes can be derived from a biological source selected from thegroup consisting of organs, tissues and cells, or they may be fromnon-natural sources as discussed in the definitions section above.Biological analytes can be selected from the group consisting of genomicDNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and a combination ofany of the foregoing. The nucleic acid array can be selected from thegroup consisting of DNA, RNA and analogs thereof, an example of thelatter being PNA. Such nucleic acids or analogs can be modified on anyone of the sugar, phosphate or base moieties. The solid support can takea number of different forms, including being porous or non-porous. Aporous solid support can be selected from the group consisting ofpolyacrylamide and agarose. A non-porous solid support may compriseglass or plastic. The solid support can also be transparent,translucent, opaque or reflective.

[0111] Nucleic acids can be directly or indirectly fixed or immobilizedto the solid support. In terms of indirect attachment, the nucleic acidscan be indirectly fixed or immobilized to the solid support by means ofa chemical linker or linkage arm.

[0112] As discussed elsewhere in this disclosure, the inherent UDT canselected from the group consisting of 3′ polyA segments, 5′ caps,secondary structures, consensus sequences and a combination of any ofthe foregoing. The consensus sequences can be selected from the groupconsisting of signal sequences for polyA addition, splicing elements,multicopy repeats and a combination of any of the foregoing. As alsodiscussed elsewhere in this disclosure, the UDEs can be selected fromthe group consisting of nucleic acids, nucleic acid analogs,polypeptides, polysaccharides, synthetic polymers and a combination ofany of the foregoing. As mentioned previously, such analogs can take theform of PNA. The UDE generates a signal directly or indirectly. Directsignal generation can take any number of forms and can be selected fromthe group consisting of a fluorescent compound, a phosphorescentcompound, a chemiluminescent compound, a chelating compound, an electrondense compound, a magnetic compound, an intercalating compound, anenergy transfer compound and a combination of any of the foregoing.Where indirect signal generation is desired, such can take a number ofdifferent forms and in this regard can be selected from the groupconsisting of an antibody, an antigen, a hapten, a receptor, a hormone,a ligand, an enzyme and a combination of any of the foregoing. Amongsuitable enzymes which can be indirectly detected, these would includeenzymes which catalyze any reaction selected from the group consistingof a fluorogenic reaction, a chromogenic reaction and a chemiluminescentreaction.

[0113] This invention also provides a composition of matter thatcomprises a library of analytes, such analytes being hybridized to anarray of nucleic acids, and such nucleic acids being fixed orimmobilized to a solid support, wherein the analytes comprise anon-inherent universal detection target (UDT) and a universal detectionelement (UDE) hybridized to the UDT, and wherein the UDE generates asignal directly or indirectly to detect the presence or quantity of suchanalytes. The nature of the analyte, the nucleic acid array,modifications, solid support are as described in the precedingparagraphs above. The non-inherent universal detection targets (UDTs)can comprise homopolymeric sequences or heteropolymeric sequences. Theuniversal detection elements (UDEs) can be selected from the groupconsisting of nucleic acids, nucleic acid analogs and modified formsthereof. The UDEs generate a signal directly or indirectly, such directand indirect signal generation also being discussed in the paragraphsjust above.

[0114] The present invention further provides a composition of matterthat comprises a library of analytes, such analytes being hybridized toan array of nucleic acids, and such nucleic acids being fixed orimmobilized to a solid support, wherein the hybridization between theanalytes and the nucleic acids generate a domain for complex formation,and the composition further comprises a signaling entity complexed tothe domain. Statements and features regarding the nature of the libraryof analytes, the nucleic acid array, the solid support and fixation orimmobilization thereto, and direct/indirect signal generation are asdiscussed hereinabove, particularly the last several paragraphs.Notably, the domain for complex formation can be selected from the groupconsisting of DNA-DNA hybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNAhybrids and RNA-PNA hybrids. The signaling entity that is complexed tothe domain can be selected from the group consisting of proteins andintercalators. Such proteins can comprise nucleic acid binding proteinswhich bind preferentially to double-stranded nucleic acid, the lattercomprising antibodies, for example. These antibodies are specific fornucleic acid hybrids and are selected from the group consisting ofDNA-DNA hybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids andRNA-PNA hybrids. In accordance with the present invention, usefulintercalators can be selected from the group consisting of ethidiumbromide, diethidium bromide, acridine orange and SYBR Green. Whenemployed in accordance with the present invention, the proteins generatea signal directly or indirectly. Such forms and manner of direct andindirect signal generation are as described elsewhere in thisdisclosure, particularly in several paragraphs above.

[0115] Related to the above described compositions are unique and usefulprocesses. The present invention thus provides a process for detectingor quantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of the nucleic acids of interest comprise at least one inherentuniversal detection target (UDT); and (iii) universal detection elements(UDE) which generates a signal directly or indirectly; b) hybridizingthe library (ii) with the array of nucleic acids (i) to form hybrids ifthe nucleic acids of interest are present; c) contacting the UDEs withthe UDTs to form a complex bound to the array; d) detecting orquantifying the more than one nucleic acid of interest by detecting ormeasuring the amount of signal generated from UDEs bound to the array.Many of these elements have been described previously in thisdisclosure, but at the risk of some redundancy, elaboration is now made.For example, the nucleic acid array can be selected from the groupconsisting of DNA, RNA and analogs thereof, the latter comprising PNA.Modifications to these nucleic acids and analogs can be usefully carriedout to any one of the sugar, phosphate or base moieties. The solidsupport can be porous, e.g., polyacrylamide and agarose, or non-porous,e.g., glass or plastic. The solid support can also be transparent,translucent, opaque or reflective.

[0116] Nucleic acids are directly or indirectly fixed or immobilized tothe solid support. Indirect fixation or immobilization to the solidsupport can be carried out by means of a chemical linker or linkage arm.As discussed elsewhere herein, the library of analytes can be derivedfrom a biological source selected from the group consisting of organs,tissues and cells, or they may be from non-natural or more synthetic orman-made sources. Among biological analytes are those selected from thegroup consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.

[0117] The inherent UDT used in the above process can be selected fromthe group consisting of 3′ polyA segments, 5′ caps, secondarystructures, consensus sequences, and a combination of any of theforegoing. Such consensus sequences can be selected from the groupconsisting of signal sequences for polyA addition, splicing elements,multicopy repeats, and a combination of any of the foregoing. UDEs canbe selected from the group consisting of nucleic acids, nucleic acidanalogs, e.g., PNA, polypeptides, polysaccharides, synthetic polymersand a combination of any of the foregoing. UDEs generate a signaldirectly or indirectly. Direct signal generation can be various and maybe selected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing. Indirect signal generation can also be various andmay be selected from the group members consisting of an antibody, anantigen, a hapten, a receptor, a hormone, a ligand, an enzyme and acombination of any of the foregoing. When desired and employed in theprocess at hand, such an enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction. Those skilled in the art will readilyappreciate that the above-described process can further comprise one ormore washing steps.

[0118] This invention provides another such process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of the nucleic acids of interest comprise at least one inherentuniversal detection target (UDT); and (iii) universal detection elements(UDE) which generates a signal directly or indirectly; b) contacting theUDEs with the UDTs in the library of nucleic acid analytes to form oneor more complexes; c) hybridizing the library of nucleic acid analyteswith the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; d) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array. The nature and form ofthe nucleic acid array, modifications, solid support, direct/indirectfixation or immobilization, library of analytes, inherent UDT, UDE,direct/indirect signal generation, and the like, are as describedelsewhere in this disclosure, including more particularly the lastseveral paragraphs above. Furthermore, this process can comprise one ormore conventional washing steps.

[0119] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of the nucleic acidsof interest comprise at least one non-inherent universal detectiontarget (UDT), wherein the non-inherent UDT is attached to the nucleicacid analytes; and (iii) universal detection elements (UDE) whichgenerate a signal directly or indirectly; b) hybridizing the library(ii) with the array of nucleic acids (i) to form hybrids if the nucleicacids of interest are present; c) contacting the UDEs with the UDTs toform a complex bound to the array; d) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array. As described variouslyin this disclosure, the nature and form of the nucleic acid array,modifications to nucleic acid and nucleic acid analogs, the solidsupport, direct and indirectfixation/immobilization to the solidsupport, the library of analytes, direct and indirect signal generation,and the like, are as described elsewhere in this disclosure. Ofparticular mention are the non-inherent universal detection targets(UDTs) which can comprise homopolymeric sequences and heteropolymericsequences. Also of particular mention are the universal detectionelements (UDEs) which can be selected from the group consisting ofnucleic acids, nucleic acid analogs, e.g., PNA, and modified formsthereof. One or more washing steps can be included in this last process.

[0120] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of such nucleic acidsof interest comprise at least one non-inherent universal detectiontarget (UDT), wherein the non-inherent UDTs are attached to the nucleicacid analytes; and (iii) universal detection elements (UDE) whichgenerate a signal directly or indirectly; b) contacting the UDEs withthe UDTs in the library of nucleic acid analytes to form one or morecomplexes; c) hybridizing the library (ii) with the array of nucleicacids (i) to form hybrids if such nucleic acids of interest are present;d) detecting or quantifying the more than one nucleic acid of interestby detecting or measuring the amount of signal generated from UDEs boundto the array. Descriptions for the nucleic acid array, modifications,solid support, direct/indirect fixation or immobilization to the solidsupport, the library of analytes, the non-inherent universal detectiontargets (UDTs), the universal detection elements (UDEs), direct/indirectsignal generation, inclusion of washing steps, and the like, are foundelsewhere in this disclosure and are equally applicable to this lastdescribed process.

[0121] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore universal detection targets (UDT) to a nucleic acid; (iv) universaldetection elements (UDE) which generates a signal directly orindirectly; b) attaching such UDTs (iii) to the library of nucleic acidanalytes (ii); c) hybridizing the library (ii) with the array of nucleicacids (i) to form hybrids if such nucleic acids of interest are present;d) contacting the UDEs with the UDTs to form a complex bound to thearray; e) detecting or quantifying the more than one nucleic acid ofinterest by detecting or measuring the amount of signal generated fromUDEs bound to the array. Many of these elements have been describedalready. These include the nucleic acid array, nucleic acid analogs,sugar, phosphate and base modifications, the solid support,direct/indirect fixation and immobilization to the solid support, thelibrary of analytes, the universal detection elements, direct/indirectsignal generation, inclusion of additional washing steps, and the like,have been described elsewhere above and below and are equally applicableto this last-mentioned process. Of special mention are attaching meanswhich add homopolymeric sequences through various enzymes, e.g., poly Apolymerase and terminal transferase. Other attaching means can be usedfor adding homopolymeric or heteropolymeric sequences, and these includeenzymatic means and enzymes selected from DNA ligase and RNA ligase.

[0122] Still another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore universal detection targets (UDT) to a nucleic acid; (iv) universaldetection elements (UDE) which generate a signal directly or indirectly;b) attaching the UDTs (iii) to the library of nucleic acid analytes(ii); c) contacting the UDEs with the UDTs in the library of nucleicacid analytes to form one or more complexes; d) hybridizing the library(ii) with the array of nucleic acids (i) to form hybrids if such nucleicacids of interest are present; e) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array. As might be expected, theelements recited in this process have been described elsewhere in thisdisclosure and are equally applicable to this last described process.These previously described elements include the nucleic acid array,modifications, the solid support, direct/indirect fixation orimmobilization to the solid support, the library of analytes, attachingmeans, UDE, direct/indirect signal generation and the inclusion ofwashing steps.

[0123] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; and (iii) universal detectionelements (UDEs) which bind to a domain formed by nucleic acid hybridsfor complex formation and generate a signal directly or indirectly; b)hybridizing the library (ii) with the array of nucleic acids (i) to formhybrids if such nucleic acids of interest are present, wherein anyformed hybrids generate a domain for complex formation; c) contactingthe UDEs with any hybrids to form a complex bound to the array; d)detecting or quantifying the more than one nucleic acid of interest bydetecting or measuring the amount of signal generated from UDEs bound tothe array. Descriptions for the nucleic acid array, nucleic acidanalogs, e.g., PNA, modifications (sugar, base and phosphate moieties),the solid support, fixation/immobilization, the library of analytes, thedomain for complex formation, direct/indirect signal generation fromsignaling proteins, washing steps, and the like, have already been givenabove and are equally applicable to this last mentioned process. Ofspecial note is this process wherein the signaling entity is complexedto the domain for complex formation, such signaling entity beingselected from proteins and intercalators. Such proteins can includenucleic acid binding proteins which bind preferentially todouble-stranded nucleic acids, e.g., antibodies, particularly suchantibodies which are specific for nucleic acid hybrids, e.g., DNA-DNAhybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids and RNA-PNAhybrids. Intercalators have also been previously described in thisdisclosure and can be selected from ethidium bromide, diethidiumbromide, acridine orange and SYBR Green.

[0124] Other compositions of matter are provided by this invention. Onesuch composition comprises a library of first nucleic acid analytecopies, such first nucleic acid copies being hybridized to an array ofnucleic acids, those nucleic acids being fixed or immobilized to a solidsupport, wherein such first nucleic acid copies comprise an inherentuniversal detection target (UDT) and a universal detection element (UDE)attached to the UDT, wherein the UDE generates a signal directly orindirectly to detect the presence or quantity of any analytes. Thelibrary of analytes, e.g., biological sources, and examples of suchanalytes, e.g., genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA,snRNA and a combination of any of the foregoing, has been describedabove. Equally so, the nucleic acid array has been already described,including, for example, DNA, RNA and analogs thereof, e.g., PNA.Modifications to the nucleic acids and analogs (sugar, phosphate, base),features of the solid support (porous/non-porous, transparent,translucent, opaque, reflective), fixation/immobilization to the solidsupport, the inherent UDT, the UDE, direct/indirect signal generationfrom UDEs have been described above and apply equally to this lastcomposition.

[0125] Another composition of matter comprises a library of firstnucleic acid analyte copies, such first nucleic acid copies beinghybridized to an array of nucleic acids, the nucleic acids being fixedor immobilized to a solid support, wherein such first nucleic acidcopies comprise one or more non-inherent universal detection targets(UDTs) and one or more universal detection elements (UDEs) attached tothe UDTs, wherein the UDEs generate a signal directly or indirectly todetect the presence or quantity of any analytes, and wherein the UDTsare either: (i) at the 5′ ends of the first nucleic acid copies and notadjacent to an oligoT segment or sequence, or (ii) at the 3′ ends of thefirst nucleic acid copies, or (iii) both (i) and (ii). The library ofanalytes, nucleic acid array, nucleic acid modifications, solid support,fixation/immobilization to the solid support, non-inherent UDTs, e.g.,heteropolymeric sequences, UDEs (e.g., nucleic acids, nucleic acidanalogs, polypeptides, polysaccharides, synthetic polymers, etc),direct/indirect signal generation from UDEs have already been describedabove and are applicable to this last described composition.

[0126] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of such nucleic acidsof interest comprise at least one inherent universal detection target(UDT); (iii) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (iv) polymerizing means for synthesizingnucleic acid copies of the nucleic acids of analytes; b) synthesizingone or more first nucleic acid copies which are complementary to all orpart of the nucleic acid analytes and synthesizing sequences which arecomplementary to all or part of the UDT to form a complementary UDT; c)hybridizing such first nucleic acid copies with the array of nucleicacids (i) to form hybrids if such nucleic acids of interest are present;d) contacting the UDEs with the complementary UDTs of the first nucleicacid copies to form a complex bound to the array; e) detecting orquantifying the more than one nucleic acid of interest by detecting ormeasuring the amount of signal generated from UDEs bound to the array.Statements and descriptions for the nucleic acid array, modifications,solid support, fixation/immobilization, the library of analytes,inherent UDTs, e.g., consensus sequences, UDEs, direct/indirect signalgeneration from UDEs, have been given above and are equally applicableto this last process. Of special mention are the recited polymerizingmeans which can be selected from E. coli DNA Pol I, Klenow fragment ofE. coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNApolymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.

[0127] Another embodiment provided by this invention is a process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical in part or whole to the nucleicacids of interest; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest sought to be detected orquantified, wherein each of such nucleic acids of interest comprise atleast one inherent universal detection target (UDT); (iii) universaldetection elements (UDE) which generate a signal directly or indirectly;and (iv) polymerizing means for synthesizing nucleic acid copies of suchnucleic acid analytes; b) synthesizing one or more first nucleic acidcopies of such nucleic acid analytes; c) contacting the UDEs with theUDTs in the first nucleic acid copies to form one or more complexes; d)hybridizing such first nucleic acid copies with the array of nucleicacids (i) to form hybrids if such nucleic acids of interest are present;and e) detecting or quantifying the more than one nucleic acid ofinterest by detecting or measuring the amount of signal generated fromUDEs bound to the array. The nucleic acid array, nucleic acidmodifications, the solid support, fixation/immobilization (direct andindirect), the library of analytes, inherent UDTs, UDEs, signalgeneration from UDEs (direct/indirect), polymerizing means, have beendescribed above. Such descriptions are equally applicable to this lastprocess.

[0128] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore non-inherent universal detection targets (UDT) to a nucleic acid;(iv) universal detection elements (UDE) which generate a signal directlyor indirectly; and (v) polymerizing means for synthesizing nucleic acidcopies of the nucleic acid analytes; b) attaching the non-inherent UDTsto either the 3′ ends of the nucleic acid analytes, the 5′ ends of thefirst nucleic acid analytes, or both the 3′ ends and the 5′ ends of thenucleic acid analytes; c) synthesizing one or more first nucleic acidcopies of the nucleic acid analytes; d) hybridizing the first nucleicacid copies with the array of nucleic acids (i) to form hybrids if suchnucleic acids of interest are present; e) contacting the UDEs with theUDTs of the first nucleic acid copies to form a complex bound to thearray; and f) detecting or quantifying the more than one nucleic acid ofinterest by detecting or measuring the amount of signal generated fromUDEs bound to the array. See many of the preceding paragraphs fordescriptions and characteristics of the nucleic acid array,modifications, the solid support, fixation/immobilization, the libraryof analytes, attaching means, UDEs, direct/indirect signal generationfrom UDEs, polymerizing means, and the like.

[0129] Yet another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore non-inherent universal detection targets (UDT) to a nucleic acid;(iv) universal detection elements (UDE) which generate a signal directlyor indirectly; and (v) polymerizing means for synthesizing nucleic acidcopies of the nucleic acid analytes; b) attaching such non-inherent UDTsto either the 3′ ends of the nucleic acid analytes, the 5′ ends of thefirst nucleic acid analytes, or both the 3′ ends and the 5′ ends of thenucleic acid analytes; c) synthesizing one or more first nucleic acidcopies of the nucleic acid analytes; d) contacting the UDEs with theUDTs of the first nucleic acid copies to form complexes; e) hybridizingthe first nucleic acid copies with the array of nucleic acids (i) toform hybrids if any nucleic acids of interest are present; f) detectingor quantifying the more than one nucleic acid of interest by detectingor measuring the amount of signal generated from UDEs bound to thearray. The nucleic acid array, modifications, the solid support,direct/indirect fixation/immobilization, the library of analytes,attachment means, UDEs, signal generation from UDEs, direct/indirectsignal generation, polymerizing means, and the like, have already beendescribed. Such descriptions are equally applicable to thislast-described process.

[0130] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to such nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore non-inherent universal detection targets (UDT) to a nucleic acid;(iv) universal detection elements (UDE) which generate a signal directlyor indirectly; and (v) polymerizing means for synthesizing nucleic acidcopies of the nucleic acid analytes; b) synthesizing one or more firstnucleic acid copies of the nucleic acid analytes; c) attaching thenon-inherent UDTs to either the 3′ ends of the first nucleic acidcopies, the 5′ ends of the first nucleic acid copies, or both the 3′ends and the 5′ ends of the first nucleic acid copies; d) hybridizingthe first nucleic acid copies with the array of nucleic acids (i) toform hybrids if any nucleic acids of interest are present; e) contactingthe UDEs with the UDTs of the first nucleic acid copies to form acomplex bound to the array; and f) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array. Descriptions for theabove-recited elements have been given above and are equally applicableto this last process.

[0131] Still another process provided by this invention is for detectingor quantifying more than one nucleic acid of interest in a librarycomprises the steps of a) providing (i) an array of fixed or immobilizednucleic acids identical in part or whole to the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)means for attaching one or more non-inherent universal detection targets(UDT) to a nucleic acid; (iv) universal detection elements (UDE) whichgenerate a signal directly or indirectly; and (v) polymerizing means forsynthesizing nucleic acid copies of the nucleic acid analytes; b)synthesizing one or more first nucleic acid copies of the nucleic acidanalytes; c) attaching the non-inherent UDTs to either the 3′ ends ofthe first nucleic acid copies, the 5′ ends of the first nucleic acidcopies, or both the 3′ ends and the 5′ ends of the first nucleic acidcopies; d) contacting the UDEs with the UDTs of the first nucleic acidcopies to form a complex; e) hybridizing the first nucleic acid copieswith the array of nucleic acids (i) to form hybrids if any nucleic acidsof interest are present; and f) detecting or quantifying the more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to the array. These elements andsubelements have been described elsewhere in this disclosure. Suchdescriptions apply to this last process.

[0132] Yet another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acidscomplementary to the nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) universal detection elements(UDEs) which bind to a domain for complex formation formed by nucleicacid hybrids and generate a signal directly or indirectly; and (iv)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes; b) synthesizing one or more nucleic acid copies of thenucleic acid analytes; c) hybridizing the first nucleic acid copies withthe array of nucleic acids (i) to form hybrids if any nucleic acids ofinterest are present, wherein any formed hybrids generate a domain forcomplex formation; d) contacting the UDEs with the hybrids to form acomplex bound to the array; and e) detecting or quantifying the morethan one nucleic acid of interest by detecting or measuring the amountof signal generated from UDEs bound to the array. The above-recitedelements and subelements and variations thereof are described elsewherein this disclosure and are equally applicable to this just-mentionedprocess.

[0133] One aspect of the present invention discloses methods thateliminate the necessity for enzymatic incorporation of labelednucleotides by an end user. In this particular aspect, common orconserved features present in a diverse population of nucleic acidanalytes are used to assay the extent of hybridization of the analytesto discrete target elements in an array format. These common orconserved features are Universal Detection Targets (UDTs) which canprovide signal generation by binding of Universal Detection Elements(UDEs).

[0134] Examples of UDTs that may be inherently present in a populationof diverse nucleic acid analytes can comprise but not be limited to 3′poly A segments, 5′ caps, secondary structures and consensus sequences.Examples of consensus sites that might find use in the present inventioncan comprise but not be limited to signal sites for poly A addition,splicing elements and multicopy repeats such as Alu sequences.

[0135] UDEs may be directly or indirectly labeled. Examples of directlylabels can comprise but not be limited to any members of a groupconsisting of a fluorescent compound, a phosphorescent compound, achemiluminescent compound, a chelating compound, an electron densecompound, a magnetic compound, an intercalating compound, an energytransfer compound and a combination of any of the foregoing.

[0136] Examples of indirect labels can comprise but not be limited toany members of a group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing. Among such enzymes are any enzymes which catalyzereactions_selected from the group consisting of a fluorogenic reaction,a chromogenic reaction and a chemiluminescent reaction.

[0137] RNA and DNA polymerases sometimes have difficulty in acceptinglabeled nucleotides as substrates for polymerization. In prior art, thisshortcoming can result in the production of a labeled library thatconsists of short strands with few signal generating entities.Limitations caused by such inefficient incorporation can be partiallycompensated for by increasing the amount of labeled precursors in thereaction mixtures. However, this method achieves only a moderateimprovement and entails a higher cost and waste of labeled reagents. Incontrast, this particular aspect of the present invention disclosesmeans by which diverse nucleic acids in a library can be hybridized inan array format in their native form without the need of anymanipulations or modifications and then be detected by the presence ofUDTs bound to the array.

[0138] An illustrative depiction of this process is given in FIG. 1.Although there are multiple unique species of mRNA that can make up adiverse population of nucleic acids in a sample, the common elementsthat are shared by these nucleic acids can be used as UDTs.Hybridization of the mRNA to an array permits the localization ofindividual species to discrete locations on the array. The determinationof the amount of sample that is bound to each locus of an array is thencarried out by detection of the amount of UDT present at each locus bybinding of the appropriate UDE. Thus, in FIG. 1, locus 1 and 3 would becapable of generating an amount of signal that would be proportionate tothe amount of mRNA bound to each of those sites. On the other hand therewould little or no signal generation from locus 2 since there was littleor no mRNA bound to that site. A single labeled species of mostly orcompletely poly T or U could be used as a UDE to quantify the amount ofpoly A tails of the various species of eucaryotic mRNA in FIG. 1. Inthis way, a single universal species of labeled material is synthesizedfor use as a UDE thereby providing an inexpensive and efficient means ofindirectly labeling the RNA molecules being quantified.

[0139] A nucleic acid UDE can be prepared either chemically orenzymatically. For example, oligonucleotide synthesizers arecommercially available that can produce a UDE consisting of labeled polyT/U sequences for detection of the poly A UDT described above. Both theamount and placement of labeled moieties can be tightly controlled bythis method. Also, since this is a homopolymeric product, probes thatare shorter by one or more bases will still be effective such that thenet yield of usable product will be higher than one that requires adiscrete specific sequence. On the other hand, methods of synthesizingsuch sequences enzymatically are also well known to those versed in theart. Commonly, a tetramer of dT is used as a primer for addition of polyT or poly U by terminal transferase. Each base can be modified to becapable of signal generation or a mixture of labeled and unlabeled basescan be used. Although A Poly A UDT has been described in the exampleabove, when different sequences are used as UDTs, the synthesis of thecorresponding UDEs can be carried out by the same chemical and enzymaticmethodologies described above. It is also contemplated that analogues ofDNA can also be used to synthesize the UDEs. For instance, instead ofusing DNA, labeled RNA or PNA (peptide nucleic acids) may also be used.

[0140] Detection and quantification of the amount of UDTs bound toparticular loci can also be carried out by the use of an antibody actingas a UDE. Examples of antibody specificities that are useful for UDEscan comprise but not be limited to recognition of the cap element at the5′ end of mature mRNAs or the homopolymeric poly A sequence.Furthermore, hybridization between nucleic acids is an event that in andof itself is capable of generating a UDT that can be recognized byantibody UDEs. For example, when a library of diverse RNA species arebound to an array, the RNA, DNA or PNA target elements in the array willgenerate RNA/RNA, RNA/DNA or RNA/PNA hybrids at each of the loci thathas homology with the particular RNA species being quantified. Althougheach of the sites has a discrete sequence, universal detection andquantification can be carried out by antibodies that recognize thechange in physical structure produced by such hybridization events.Alternatively, the hybridization between a UDE and the complementary UDTof a nucleic acid bound to the target elements of the array can bedetected by an appropriate antibody. The antibodies that are specificfor the UDEs described above can be labeled themselves or secondarylabeled antibodies can be used to enhance the signal.

[0141] If only a single library of mRNA is being analyzed, binding of aUDE to a UDT may take place before or after hybridization of the RNA toan array of detection probes. The particular order of events will dependupon the nature and stability of the binding partners. When analytesfrom two libraries are intended to be compared simultaneously, bindingof each UDE to a binding partner is preferably carried out prior tohybridization of the RNA to an array of target elements such that eachlibrary is differentially labeled. Although comparisons are typicallycarried out between two libraries, any number of comparisons can be madesimultaneously as long as each library is capable of generating a signalthat can be distinguished from the other libraries. On the other hand,rather than simultaneous hybridization and detection, the arrays can beused in a parallel or sequential fashion. In this format, hybridizationand detection is carried out separately for each library and theanalysis of the results is compared afterwards relative to normalizedcontrols of steady state genes.

[0142] In another aspect of the present invention, UDTs or UDEs areartificially incorporated into the diverse nucleic acids of the library.Enzymes that find particular use with RNA analytes may comprise but notbe limited to Poly A polymerase which specifically adds Adenineribonucleotides to the 3′ end of RNA and RNA ligase which can add anoligonucleotide or polynucleotide to either the 5′ or 3′ end of an RNAanalyte. By these means, either homopolymeric or unique sequences can beadded to serve as UDTs or UDEs. Enzymes that find particular use withDNA analytes may comprise but not be limited to Terminal Transferase foraddition to 3′ ends and DNA ligase for addition to either 3′ or 5′ ends.The sequences that are introduced into the nucleic acid analytes can belabeled during synthesis or addition of a UDE or conversely unlabeledUDTs can be synthesized or added that are detected later bycorresponding labeled UDEs. This aspect enjoys special utility whenunspliced RNA, snRNA, or rRNA are used as analytes since they may belacking inherent elements that are present in mRNA that have previouslycited as being useful as UDTs. This aspect of the present invention willalso find use with procaryotic mRNA since the poly A additions, 5′ capsand splicing elements which have been previously cited as potential UDTsof mRNA are intrinsically lacking in procaryotes.

[0143] This particular aspect of the present invention may also be usedin conjunction with fragmentation processes. For instance, mRNAmolecules from eucaryotic organisms can be very large even afterprocessing events have taken place. This size factor can hinderhybridization or allow scissions between the segment used for binding toa target element in the array and the UDT that is being used for signalgeneration. Additionally, a fragmentation step may also reduce theamount of secondary structure present in RNA. Therefore, in this aspectof the present invention, RNA can be fragmented into smaller sizedpieces either by physical or enzymatic followed by addition of sequencesthat can act as UDTs or UDEs. Examples of physical means forfragmentation of nucleic acids can include but not be limited toshearing or alkali treatment. Examples of enzymatic means can includebut not be limited to a partial nuclease or RNase digestion.

[0144] In addition, DNA from most sources will also be extremely largein its native form. DNA analytes may also be fragmented by suitablephysical or enzymatic means. A particularly useful enzymatic means wouldbe the use of restriction enzymes where the nature of the recognitionsequence for the restriction enzyme will determine the average size ofthe fragments. Also, although most restriction enzymes requiredouble-stranded DNA as templates, some enzymes such as Hha I, Hin P1 Iand MnI I cleave single-stranded DNA efficiently (2000-2001 catalog, NewEngland BioLabs, Beverly, Mass., p214). By this fragmentation method asingle analyte molecule is converted into multiple subfragments that caneach have their own artificially introduced UDT or UDE. An exemplaryillustration of this particular aspect of the present invention isincluded in FIG. 2.

[0145] In another aspect of the present invention, the diverse nucleicacids in a library are used as templates for synthesis of complementarynucleic acid copies instead of using the analytes directly for arrayanalysis. The analyte templates may have intrinsic UDTs present or theymay have UDTs artificially incorporated by the means cited earlier. Onthe other hand, the UDTs do not have to be present in the analytetemplates and incorporation of artificial UDTs can take place eitherduring or after synthesis of nucleic acid copies. Examples of enzymesthat may be used for making copies of DNA templates can comprise but notbe limited to DNA polymerases for synthesis of DNA copies and RNApolymerases for the synthesis of RNA copies. Examples of DNA polymerasesthat may have use in the present invention for synthesis of DNA copiesfrom DNA templates can include but not be limited to E.coli DNA Pol I,the Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA polymerase, T4 DNA polymerase,T7 DNA polymerase, ALV Reverse Transcriptase, RSV Reverse Transcriptase,HIV-1 Reverse Transcriptase, HIV-2 Reverse Transcriptase, Sensiscript,Omniscript and various mutated or otherwise altered forms of theforegoing. Examples of RNA polymerases that may have use in the presentinvention for synthesis of RNA copies from DNA templates can include butnot be limited to bacteriophage T3 RNA polymerase, bacteriophage T7 RNApolymerase and bacteriophage SP6 RNA polymerase. Examples of enzymesthat may have use in the present invention for making DNA copies of RNAtemplates can comprise but not be limited to ALV Reverse Transcriptase,RSV Reverse Transcriptase, HIV-1 Reverse Transcriptase, HIV-2 ReverseTranscriptase, Sensiscript, Omniscript, Bst DNA polymerase, Bca DNApolymerase, Tth DNA polymerase and various mutated or otherwise alteredforms of the foregoing.

[0146] Examples of enzymes that may have use in the present inventionfor making RNA copies of RNA templates can comprise but not be RNAdependent RNA polymerases (Koonin, 1991 J. Gen Virol. 72; 2197-2206,incorporated herein by reference).

[0147] Efficient synthesis of complementary copies of analyte templatesrequire the presence of a promoter for efficient synthesis by DNAdependent RNA polymerases while the other cited exemplary enzymesrequire primers. Incorporation of a UDT into a DNA analyte that will betranscribed by a DNA dependent RNA polymerase can comprise but not belimited to ligation of a UDT sequence and a promoter sequence by theaction of DNA ligase. This process is depicted below:

DNA analyte+UDT-Promoter=DNA Analyte-UDT-Promoter

[0148] Transcription of this construct would then be capable ofproduction of RNA with the structure: 3′ analyte-DT 5′:

[0149] One means of carrying out this particular aspect of the presentinvention is digestion of a library of diverse double-stranded DNAanalytes by a restriction enzyme followed by ligation of adouble-stranded DNA segment comprising an RNA promoter sequence.Subsequent transcription of the transcription units can synthesizeeither labeled or unlabeled transcripts. The unlabeled transcripts canbe detected by the presence of either inherent or synthetically addedUDTs.

[0150] When primers are used for synthesis of complementary copies ofanalyte templates, the primers can comprise random sequences or selectedsequences for binding to the analyte templates. Random primers that havecommonly been used for priming events have ranged from hexamers tododecamers. Selected sequences that are useful as primers can becomplementary to inherent sequences or to non-inherent sequences thathave been introduced into the analyte templates. Examples of inherentsequences can include but not be limited to consensus sequences orhomopolymeric sequences. Consensus sequences can be derived fromelements that are retained in a large portion of the population beingstudied. Examples of these could comprise but not be limited to poly Aaddition sites, splicing elements and multicopy repeats such as Alusequences. An example of inherent homopolymeric sequences used forprimer binding can be the poly A tail that is intrinsic to mature mRNAin eucaryotes. Non-inherent homopolymeric or unique sequences that canbe used for primer binding may be introduced into RNA templates by meansthat can include but not be limited to poly A polymerase or RNA ligase.Non-inherent homopolymeric or unique sequences that can be used forprimer binding may be introduced into DNA templates by means that caninclude but not be limited to Terminal Transferase and DNA ligase. Theartificial binding sites can be introduced into intact nucleic acidtemplates or fragmentation processes may be carried out as describedpreviously.

[0151] When homopolymeric or conserved sequences are used as primerbinding sites, the library can be subdivided by the use of primers thathave been synthesized with 1 or more additional discrete bases at the 3′end. For example, an oligonucleotide primer that has the formula5′-T_(n)dC-3′ would preferentially prime mRNAs whose last base was a Gbefore the poly A tail rather than priming the entire population ofmRNA's with poly A tails. The same principle would also hold true wheneither 5′-T_(n)dG-3′ or 5′-T_(n)dA-3′ primers are used. This wouldprovide three separate sub-populations of copies of the original mRNApopulation that in toto should encompass the entire RNA population withpoly A tails. This population could be further divided by inclusion of a2^(nd) discrete base at the 3′ end of the primers. In this case,oligonucleotides would have either dC, dG, dA or dT as the last base atthe 3′ end and dC, dG or dA in the penultimate position and theremaining portion comprising a poly T segment. This would create thepotential for 12 separate pools from the original population. Furtherprovision of discrete bases at the 3^(rd) nucleotide position from the3′ end would provide a separation into 48 different subpopulations ifdesired and so on.

[0152] The use of subpopulations may have utility in providing RNA withlower complexity thereby simplifying analysis later on. In addition, theuse of discrete bases at the 3′ end would limit the size of poly T tailsat the end of the cDNA copies since significant amounts of primingevents will only take place at the junction of the poly A addition site.This may reduce background hybridization caused by extensive polyT orPolyA tracts. Also it may increase yields of labeled products bydecreasing stalling or premature terminations caused by longhomopolymeric tracts. On the other hand, the use of a mixture of oligo Tprimers with discrete bases at the 3′ end would be similar to acompletely homopolymeric oligo T primer in being able to synthesize acomplete representation of the original analyte sequences while at thesame retaining the ability to constrain the size of homopolymeric tails.

[0153] In this particular aspect of the present invention, the cDNAmolecules synthesized from the pool of RNA templates also comprise UDTsor UDEs. As described previously, these UDTs can be inherently presentor they may be non-inherent sequences that are artificially incorporatedduring synthesis of cDNA. When an analyte has a nucleic acid sequencethat can be used as a UDT, synthesis of the complementary copy creates asequence that can also be used as a UDT. For example, the poly Asequence at the 3′ end of eucaryotic mRNA was previously described as apotential UDT. When this mRNA is used as a template by extension of apoly T primer with or without additional bases, the poly T segment ofthe cDNA copy can function as a UDT. The destruction or separation ofthe RNA templates from the cDNA would allow the poly T at the 5′ end ofthe cDNA to act as a UDT by binding of a labeled poly A UDE. UDTs orUDEs can also be incorporated into cDNA copies by inclusion of nucleicacid segments that don't participate in primer binding into the 5′ tailsof either random, homopolymeric, or specific sequence primers. Theparticular sequence of the additional nucleic acid segments used as UDTsare of arbitrary nature since they aren't needed for primer binding. Assuch, the choice of sequence for these UDTs can range in complexity fromhomopolymeric sequences to specific unique sequences. Their nature isalso arbitrary, and either the primer or the UDT can comprise PNA's orother nucleic acid homologues. In addition, they may be other polymericentities besides nucleic acids that provide recognition for UDEs.

[0154] Since the nature of the UDT or UDE can be selected by the user,the present invention allows simple differentiation between librariesthat are being compared. For instance, one population that is beingstudied can be extended by homopolymeric or random primers andhybridized with a UDE labeled with Cy 3. A second population that isbeing compared can be extended by homopolymeric or random primers andhybridized with UDEs that have Cy 5 incorporated into them. The otherend of the cDNA is also available for use with UDEs. For example, aftersynthesis of cDNA copies by reverse transcriptase, the 3′ ends can beextended further by the non-template directed addition of nucleotides byTerminal Transferase. An illustration of this particular aspect of thepresent invention is included in FIG. 3.

[0155] Detection of the presence of UDTs or UDEs in the library orlibraries of various nucleic acids can be carried out by any of themeans that have been described previously for UDTs. If only a singlelibrary is being analyzed, binding of a probe or antibody to a 5′ or 3′UDT or UDE may take place before or after hybridization of nucleic acidsto the detection elements of the array. The particular order of eventswill depend upon the nature and stability of the binding partners. Onthe other hand, when each population incorporates a different UDT orUDE, binding of labeled moieties to the UDTs can take place eitherbefore or after hybridization of the copies of the analyte to an array.However, as described previously, the same UDT or UDE can be used foreach population if parallel or sequential hybridizations are carriedout.

[0156] It is also contemplated that the various aspects of the presentinvention can be used to augment rather than substitute for otherpreviously disclosed methods. For instance, signal can be generated incDNA copies by a labeled primer being extended in the presence oflabeled nucleotides. The signal generated by such a method would be asummation of the signal generated by the original primer and whateverlabeled nucleotides were incorporated during strand extension. Thus, acombination of methodologies would generate a signal that would behigher than the amount that would be achieved by either method alone. Inaddition to a pre-labeled primer, the other methods that are disclosedin the present invention can also be used in various combinations.

[0157] There may be situations where amplification of sequences in asample is advantageous. Therefore, in another aspect of the presentinvention, multiple cycles of synthesis can be carried out to generatelinear amplification of a library of diverse nucleic acid sequences. Inthe first step of this particular aspect of the present invention, theentire population or a subset of the population of nucleic acidsanalytes is used to synthesize 1^(st) strand nucleic acid copies.Whether the initial analyte is DNA or RNA, in the context of the presentinvention, this product is considered to be a cNA since it represents anucleic acid copy of the analyte. Synthesis of the 1^(st) strand nucleicacid copies can be carried out as described previously by using discreteprimers, random primers, homopolymers, or homopolymers with one or morediscrete bases at their 3′ ends. In this particular embodiment of thepresent invention, priming with homopolymers with one or more discretebases at their 3′ ends may also increase the efficiency of amplificationsince resources such as primers and substrates will be directed onlytowards amplification of a discrete subpopulation derived from the1^(st) cNA synthesis reaction.

[0158] For linear amplification, a primer binding site on a nucleic acidanalyte is used multiple times by separation of a 1^(st) cNA copy fromits template followed by reinitiation of a new 1^(st) cNA copy.Separation can be carried out by exposure of the reaction mix to hightemperature. If the enzyme used for nucleic acid synthesis is Taqpolymerase, Tth polymerase or some other heat stable polymerase themultiple reactions can be carried out by thermocycling of the reactionwithout the addition of any other reactions. On the other hand, if highdenaturation temperatures are used in conjunction with enzymes that areheat labile, for instance Bst DNA polymerase, Klenow fragment of Pol Ior MuLV Reverse Transcriptase, irreversible heat inactivation of theenzyme takes place and the enzyme has to be replenished for furtherrounds of cNA synthesis. Alternatively, methods have been disclosed byFuller in U.S. Pat. No. 5,432065 and by Lakobashvill and Lapidot, 1999(Nucleic Acids Research 27; 1566-1568) for reagents that allow lowtemperature denaturation of nucleic acids for use with PCR, both ofwhich methods are incorporated by reference. Furthermore, Winhoven andRossau have disclosed in PCT Application WO 98/45474 (also incorporatedby reference) that temperature manipulation can be avoided completely byelectrically controlled manipulation of divalent ion levels. Thus bythese methods even thermo-labile enzymes can carry out multiple cyclesof synthesis for linear amplification. Both above-cited patent documentsand the above-cited publication are incorporated herein by reference.

[0159] Amplification is a significant aspect of this invention. Severalcompositions and processes are devoted and directed to amplification.For example, provided herein is a composition of matter comprising alibrary of double-stranded nucleic acids substantially incapable of invivo replication and free of non-inherent homopolymeric sequences, thenucleic acids comprising sequences complementary or identical in part orwhole to inherent sequences of a library obtained from a sample, whereinthe double-stranded nucleic acids comprise at least one inherentuniversal detection target (UDT) proximate to one end of the doublestrand and at least one non-inherent production center proximate to theother end of the double strand. The sample from which the inherentsequences of the library are obtained can comprise biological sources,e.g., organs, tissues and cells. As described elsewhere herein, thelibrary of nucleic acids can be derived from genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing. Inherent UDTs can be selected from the group consisting of 3′polyA segments, consensus sequences, or both. As already describedabove, consensus sequences can be selected from the group consisting ofsignal sequences for poly A addition, splicing elements, multicopyrepeats, and a combination of any of the foregoing. Of special mentionis the production center which can be selected from the group consistingof primer binding sites, RNA promoters, or a combination of both. SuchRNA promoters can comprise phage promoters, e.g., T3, T7 and SP6.

[0160] Another composition of matter for amplification purposescomprises a library of double-stranded nucleic acids substantiallyincapable of in vivo replication, such nucleic acids comprisingsequences complementary or identical in part or whole to inherentsequences of a library obtained from a sample, wherein thedouble-stranded nucleic acids comprise at least four (4) non-inherentnucleotides proximate to one end of the double strand and a non-inherentproduction center proximate to the other end of the double strand.Descriptions for such elements, i.e., the sample, the library of nucleicacids, inherent UDTs, non-inherent nucleotides, non-inherent productioncenters, e.g., RNA promoters, e.g., phage promoters (T3, T7 and SP6) aregiven elsewhere in this disclosure and are equally applicable to thislast composition.

[0161] Another composition of matter for amplification comprises alibrary of double-stranded nucleic acids fixed to a solid support, thosenucleic acids comprising sequences complementary or identical in part orwhole to inherent sequences of a library obtained from a sample and thenucleic acids further comprising at least one first sequence segment ofnon-inherent nucleotides proximate to one end of the double strand andat least one second sequence segment proximate to the other end of thedouble strand, the second sequence segment comprising at least oneproduction center. Of special mention is the use of beads as the solidsupport, particularly beads and magnetic beads. Other elements, such asthe sample and biological sources, the library of nucleic acids,inherent UDTs, non-inherent production centers, have already beendescribed.

[0162] Yet another amplification type composition of matter comprises alibrary of double-stranded nucleic acids attached to a solid support,the nucleic acids comprising sequences complementary or identical inpart or whole to inherent sequences of a library obtained from a sample,wherein the double-stranded nucleic acids comprise at least one inherentuniversal detection target (UDT) proximate to one end of the doublestrand and at least one non-inherent production center proximate to theother end of the double strand. The elements and subelements (solidsupport, beads, magnetic beads, sample, library of nucleic acids,inherent UDTs, consensus sequences, production centers, RNA promoters,phage promoters, e.g., T3, T7 and SP6, have been described above.

[0163] Among useful processes for detecting or quantifying more than onenucleic acid of interest in a library, one such process of the presentinvention comprises the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, the polymerizing meanscomprising a first set of primers and a second set of primers, whereinthe second set of primers comprises at least two segments, the firstsegment at the 3′ end comprising random sequences, and the secondsegment comprising at least one production center; (iv) means forsynthesizing nucleic acid copies under isothermal or isostaticconditions; b) contacting the library of nucleic acid analytes with thefirst set of primers to form more than one first bound entity; c)extending the bound first set of primers by means of template sequencesprovided by the nucleic acid analytes to form first copies of theanalytes; d) contacting the extended first copies with the second set ofprimers to form more than one second bound entity; e) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; f)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; g) hybridizing any nucleic acid copies formed instep f) to the array of nucleic acids provided in step a) (i); and h)detecting or quantifying any of the hybridized copies obtained in stepg). Elements recited in the process just above and their subelementshave already been described in this disclosure. Of special mention isthe first set of primers which are complementary to inherent UDTs.Further mention should be made that the hybridized nucleic acids cancomprise one or more signaling entities attached or incorporatedthereto. As described variously above, signal detection can be carriedout directly or indirectly. Mention is also made that the process canfurther comprise the step of separating the first copies obtained fromstep c) from their templates and repeating step b). Other steps can alsobe included such as the step of separating the extended second set ofprimers obtained from step f) from their templates and repeating stepe). Step g) can also be carried out repeatedly, a feature provided bythis invention and this last-described process. Further, means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification. These are all contemplated for use ofthis process.

[0164] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set of primersand a second set of primers, wherein the first set of primers compriseat least one production center; and (iv) means for synthesizing nucleicacid copies under isothermal or isostatic conditions; b) contacting thelibrary of nucleic acid analytes with the first set of primers to formmore than one first bound entity; c) extending the bound first set ofprimers by means of template sequences provided by the nucleic acidanalytes to form first copies of the analytes; d) extending the firstcopies by means of at least four (4) or more non-inherent homopolymericnucleotides; e) contacting the extended first copies with the second setof primers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; g)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph). Of special mention is the use or addition of terminal transferase inor after extending step d) wherein the four or more non-inherenthomopolymeric nucleotides are themselves added. Elements and subelementsof this process are described above. Special mention is made of certainaspects of this process. For example, means for synthesizing nucleicacid copies under isothermal or isostatic conditions can be carried outby one or more members selected from the group consisting of RNAtranscription, strand displacement amplification and secondary structureamplification. Moreover, the step of separating the first copiesobtained from step c) from their templates and repeating step b) can beadded to this process. Moreover, the extended second set of primersobtained from step f) can be separated from their templates and thenstep e) can be repeated as necessary or desired. In fact, step g) can berepeated as often as desired or deemed necessary.

[0165] A process for detecting or quantifying more than one nucleic acidof interest in a library comprises the steps of a) providing (i) anarray of fixed or immobilized nucleic acids identical or complementaryin part or whole to sequences of the nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) polymerizing meansfor synthesizing nucleic acid copies of the nucleic acid analytes, suchpolymerizing means comprising a first set of primers and a second set ofprimers, wherein the first set comprises at least one production center;(iv) a set of oligonucleotides or polynucleotides complementary to atleast one segment or sequence of the second set of primers; and(v) meansfor ligating the set of oligonucleotides or polynucleotides (iv); b)contacting the library of nucleic acid analytes with the first set ofprimers to form more than one first bound entity; c) extending the boundfirst set of primers by means of template sequences provided by thenucleic acid analytes to form first copies of the analytes; d) ligatingthe set of oligonucleotides or polynucleotides a) (iv) to the 3′ end ofthe first copies formed in step c) to form more than one ligatedproduct; e) contacting the ligated product with the second set ofprimers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe ligated products formed in step d) to form more than one complexcomprising the ligated products and the extended second set of primers;g) synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph). Aspects of this process, including the nucleic acid array,modifications, solid support, fixation/immobilization, nucleic acidanalytes, UDTs, production centers, signal generation, polymerizingmeans, additional steps and repeating steps, synthesizing means, and soforth, have been described above and apply equally to thislast-mentioned process. Of special mention are the above-recitedligating means which can comprise, for example, T4 DNA ligase.

[0166] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set of primersand a second set of primers, wherein the second set comprises at leastone production center; (iv) a set of oligonucleotides or polynucleotidescomplementary to at least one segment or sequence of the second set ofprimers; and (v) means for ligating the set of oligonucleotides orpolynucleotides (iv); b) contacting the library of nucleic acid analyteswith the first set of primers to form more than one first bound entity;c) extending the bound first set of primers by means of templatesequences provided by the nucleic acid analytes to form first copies ofthe analytes; d) ligating the set of oligonucleotides or polynucleotidesa) (iv) to the 3′ end of the first copies formed in step c) to form morethan one ligated product; e) contacting the ligated product with thesecond set of primers to form more than one second bound entity; f)extending the bound second set of primers by means of template sequencesprovided by the ligated products formed in step d) to form more than onecomplex comprising the ligated products and the extended second set ofprimers; g) synthesizing from a production center in the second set ofprimers in the complexes one or more nucleic acid copies underisothermal or isostatic conditions; h) hybridizing the nucleic acidcopies formed in step g) to the array of nucleic acids provided in stepa) (i); and i) detecting or quantifying any of the hybridized copiesobtained in step h). Each of the above-recited elements in this processhave been described elsewhere in this disclosure. Such descriptions areequally applicable to this process. Of special mention is the processwherein the first set of primers comprise one or more sequences whichare complementary to inherent UDTs. The hybridized nucleic acid copiescan further comprise one or more signaling entities attached orincorporate thereto. If so, previously described embodiments for signalgeneration and detection, e.g., direct and indirect generation anddetection, are applicable to this process. As described previously forother similar processes, additional steps can be carried out. Forexample, the step of separating the first copies obtained from step c)from their templates and then repeating step b) can be carried out. Afurther step of separating the extended second set of primers obtainedfrom step f) from their templates and then repeating step e) can becarried out. Also, step g) can be carried out repeatedly.

[0167] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; and (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set ofprimers, a second set of primers and a third set of primers wherein thethird set comprises at least one production center; and b) contactingthe library of nucleic acid analytes with the first set of primers toform a first set of bound primers; c) extending the first set of boundprimers by means of template sequences provided by the nucleic acidanalytes to form first copies of the analytes; d) contacting theextended first copies with the second set of primers to form a secondset of bound primers; e) extending the second set of bound primers bymeans of template sequences provided by the extended first copies toform second copies of the nucleic acid analytes; f) contacting thesecond copies with the third set of primers to form more than one thirdbound entity to form a third set of bound primers; g) extending thethird set of bound primers by means of template sequences provided bythe extended second set of primers to form a hybrid comprising a secondcopy, a third copy and at least one production center; h) synthesizingfrom the production center in the second set of primers in the complexesone or more nucleic acid copies under isothermal or isostaticconditions; i) hybridizing the nucleic acid copies formed in step i) tothe array of nucleic acids provided in step a) (i); and j) detecting orquantifying any of the hybridized copies obtained in step i). Elementsrecited in this process and variations and subelements are as describedelsewhere in this disclosure. Of special mention is the use of randomprimers as the second set of primers. Furthermore, the second set ofprimers can be complementary to the primer binding site where theprocess comprises an additional step c′) of including a primer bindingsite after carrying out step c). The primer binding site can be added bymeans of T4 DNA ligase or terminal transferase. Other aspects orvariations of this process can be made or carried out. The further stepof separating the extended second set of primers obtained from step f)from their templates and then repeating step e) can be made. Step g) canalso be carried out repeatedly. An additional step f′) of separating theextended second set of primers obtained in step e) can be carried out.Also, the step of separating the first copies obtained from step c) fromtheir templates and then repeating step b) can be carried out. Further,the step of separating the extended second set of primers obtained fromstep f) from their templates and then repeating step e) can be carriedout. Step g) can also be carried out repeatedly. In another variation ofthis process, the second set of primers can comprise at least oneproduction center which differs in nucleotide sequence from theproduction center in the third set of primers.

[0168] Still another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; and (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set of primersand a second set of primers, wherein the first set of primers are fixedor immobilized to a solid support, and wherein the second set of primerscomprises at least two segments, the first segment at the 3′ endcomprising random sequences, and the second segment comprising at leastone production center; (iv) means for synthesizing nucleic acid copiesunder isothermal or isostatic conditions; b) contacting the library ofnucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) contacting the extended firstcopies with the second set of primers to form more than one second boundentity; e) extending the bound second set of primers by means oftemplate sequences provided by the extended first copies to form morethan one complex comprising extended first copies and extended secondset of primers; f) synthesizing from a production center in the secondset of primers in the complexes one or more nucleic acid copies underisothermal or isostatic conditions; g) hybridizing the nucleic acidcopies formed in step f) to the array of nucleic acids provided in stepa) (i); and h) detecting or quantifying any of the hybridized copiesobtained in step g). The above-recited elements and variations andsubelements thereof have been described elsewhere and previously in thisdisclosure. Those descriptions apply equally to this process.

[0169] Another significant process worth discussion is one for detectingor quantifying more than one nucleic acid of interest in a library. Thisprocess comprises the steps of a) providing (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of the nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of the nucleic acid analytes, such polymerizingmeans comprising a first set of primers and a second set of primers,wherein the first set of primers are fixed or immobilized to a solidsupport, and wherein the first set of primers comprise at least oneproduction center; and (iv) means for synthesizing nucleic acid copiesunder isothermal or isostatic conditions; b) contacting the library ofnucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) extending the first copies bymeans of at least four (4) or more non-inherent homopolymericnucleotides; e) contacting the extended first copies with the second setof primers to form more than one second bound entity; f) extending thebound second set of primers by means of template sequences provided bythe extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; g)synthesizing from a production center in the second set of primers inthe complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing the nucleic acid copies formed instep g) to the array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of the hybridized copies obtained in steph). The elements recited above in this process and variations andsubelements are described elsewhere in this disclosure. Thosedescriptions apply to this process.

[0170] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set of primersand a second set of primers, wherein the first set of primers are fixedor immobilized to a solid support, and wherein the first set comprisesat least one production center; (iv) a set of oligonucleotides orpolynucleotides complementary to at least one segment or sequence of thesecond set of primers; and (v) means for ligating the set ofoligonucleotides or polynucleotides (iv); b) contacting the library ofnucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) ligating the set ofoligonucleotides or polynucleotides a) (iv) to the 3′ end of the firstcopies formed in step c) to form more than one ligated product; e)contacting the ligated product with the second set of primers to formmore than one second bound entity; f) extending the bound second set ofprimers by means of template sequences provided by the ligated productsformed in step d) to form more than one complex comprising the ligatedproducts and the extended second set of primers; g) synthesizing from aproduction center in the second set of primers in the complexes one ormore nucleic acid copies under isothermal or isostatic conditions; h)hybridizing the nucleic acid copies formed in step g) to the array ofnucleic acids provided in step a) (i); and i) detecting or quantifyingany of the hybridized copies obtained in step h). Descriptions for anyof the above-recited elements in this process are given elsewhere inthis disclosure, and need not be repeated except to say that such areequally applicable to this process.

[0171] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set of primersand a second set of primers, wherein the first set of primers are fixedor immobilized to a solid support, and wherein the second set comprisesat least one production center; (iv) a set of oligonucleotides orpolynucleotides complementary to at least one segment or sequence of thesecond set of primers; and (v) means for ligating the set ofoligonucleotides or polynucleotides (iv); b) contacting the library ofnucleic acid analytes with the first set of primers to form more thanone first bound entity; c) extending the bound first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first copies of the analytes; d) ligating the set ofoligonucleotides or polynucleotides a) (iv) to the 3′ end of the firstcopies formed in step c) to form more than one ligated product; e)contacting the ligated product with the second set of primers to formmore than one second bound entity; f) extending the bound second set ofprimers by means of template sequences provided by the ligated productsformed in step d) to form more than one complex comprising the ligatedproducts and the extended second set of primers; g) synthesizing from aproduction center in the second set of primers in the complexes one ormore nucleic acid copies under isothermal or isostatic conditions; h)hybridizing the nucleic acid copies formed in step g) to the array ofnucleic acids provided in step a) (i); and i) detecting or quantifyingany of the hybridized copies obtained in step h). For a description ofthe elements recited in this process, refer to any of the severalpreceding paragraphs.

[0172] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of the nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; and (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacid analytes, such polymerizing means comprising a first set ofprimers, a second set of primers and a third set of primers, wherein thefirst set of primers are fixed or immobilized to a solid support, andwherein the third set comprises at least one production center; and b)contacting the library of nucleic acid analytes with the first set ofprimers to form more than one first bound entity; c) extending the boundfirst set of primers by means of template sequences provided by thenucleic acid analytes to form first copies of the analytes; d)contacting the extended first copies with the second set of primers toform more than one second bound entity; e) extending the bound secondset of primers by means of template sequences provided by the extendedfirst copies to form an extended second set of primers; f) separatingthe extended second set of primers obtained in step e); g) contactingthe extended second set of primers with the third set of primers to formmore than one third bound entity; h) extending the third bound entity bymeans of template sequences provided by the extended second set ofprimers to form more than one complex comprising the extended thirdbound entity and the extended set of primers; i) synthesizing from aproduction center in the second set of primers in the complexes one ormore nucleic acid copies under isothermal or isostatic conditions; j)hybridizing the nucleic acid copies formed in step i) to the array ofnucleic acids provided in step a) (i); and k) detecting or quantifyingany of the hybridized copies obtained in step j). See this disclosurefor a discussion of any of the above-recited elements.

[0173] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to sequences of the nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; and (iii) polymerizingmeans for synthesizing nucleic acid copies of the nucleic acid analytes,such polymerizing means comprising a first set of primers; b) contactingthe nucleic acid analytes with the first set of primers to form a firstbound entity; c) extending the bound set of first set of primers bymeans of template sequences provided by the nucleic acid analytes toform first nucleic acid copies of the analytes; d) separating the firstnucleic acid copies from the analytes; e) repeating steps b), c) and d)until a desirable amount of first nucleic acid copies have beensynthesized; f) hybridizing the nucleic nucleic acid copies formed instep e) to the array of nucleic acids provided in step (i); and g)detecting or quantifying any of the hybridized first nucleic acid copiesobtained in step f). These elements are described elsewhere in thisdisclosure.

[0174] Another process for detecting or quantifying more than onenucleic acid of interest in a library comprises the steps of a)providing (i) an array of fixed or immobilized nucleic acids identicalin part or whole to sequences of the nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) polymerizing meansfor synthesizing nucleic acid copies of the nucleic acid analytes, suchpolymerizing means comprising a first set of primers and a second set ofprimers; (iv) means for addition of sequences to the 3′ end of nucleicacids; b) contacting the nucleic acid analytes with the first set ofprimer to form a first bound entity; c) extending the bound set of firstset of primers by means of template sequences provided by the nucleicacid analytes to form first nucleic acid copies of the analytes; d)extending the first nucleic copies by the addition of non-templatederived sequences to the 3′ end of the first nucleic acid copies; e)contacting the extended first nucleic acid copies with the second set ofprimers to form a second bound entity; f) extending the bound set ofsecond set of primers by means of template sequences provided by theextended first nucleic acid copies to form second nucleic acid copies;g) separating the second nucleic acid copies from the extended firstnucleic acid copies; h) repeating steps e), f) and g) until a desirableamount of second nucleic acid copies have been synthesized; i)hybridizing the second nucleic acid copies formed in step h) to thearray of nucleic acids provided in step (i); and j) detecting orquantifying any of the hybridized second nucleic acid copies obtained instep i). Descriptions for any of the above-recited elements are providedelsewhere in this disclosure.

[0175] An illustrative example of this aspect of the present inventionwould be to bind a poly T primer to poly A mRNA and extend it by Tth DNApolymerase under conditions that allow it to be used as a ReverseTranscriptase. Thermal denaturation followed by binding of an unextendedpoly T primer would allow synthesis of another copy by Tth DNAPolymerase. The amount of amplification would be proportional to a) thenumber of primer binding sites on an individual template molecule b) theefficiency of binding/extension and c) the number of cycles carried out.Thus, with a single primer binding site in a target analyte, 50%efficiency and 100 cycles denaturation/repriming, the method of thepresent invention can produce 50 1^(st) cNA copies from a single analytemolecule.

[0176] In another aspect of the present invention, primers are used togenerate a library of nucleic acids with production centers capable ofsynthesizing multiple nucleic acid copies that comprise sequences thatare either identical or complimentary to sequences in the originalanalytes. In the first step of this particular aspect of the presentinvention, the entire population or a subset of the population ofnucleic acids analytes is used to synthesize 1^(st) strand nucleic acidcopies as described previously for linear amplification. In the nextstep of this aspect of the present invention, the 1^(st) cNA strand ismade available for further binding/extension events by the removal ordestruction of the template strands. This can be carried out by avariety of physical, chemical and enzymatic means. Examples of suchmethods can consist of but not be limited to denaturation, alkali orRNase treatments. Denaturation can be carried out by exposure to highheat or by the other methods described above for multiple cycles oflinear amplification, thereby allowing them to participate in latersteps. In the next step, primers are annealed to the 1^(st) cNA strandin order to synthesize the complementary strands, thereby generatingdouble-stranded cNA copies of the original analyte population. Theprimers used for 2^(nd) strand synthesis are designed such that their 5′ends comprise sequences capable of acting as production centers. Adescription of such production centers is disclosed in Rabbani et al.,U.S. patent application Ser. No. 08/574,443, filed on Dec. 15, 1995(Novel Property Effecting And/Or Property Exhibiting Compositions forTherapeutic and Diagnostic Uses), abandoned in favor of U.S. patentapplication Ser. No. 08/978,632, filed on Nov. 25, 1997), incorporatedherein by reference. An example of a production center that would beparticularly useful in the present invention would comprise an RNApromoter segment.

[0177] For example, random hexamer primers for 2^(nd) strand synthesiscan have the structure:

[0178] 5′-promoter-N₁N₂N₃N₄N₅N₆-3′

[0179] In a preferred mode, the promoter is a phage promoter. Thesequences specific for their cognate polymerases are sufficiently shortthat their addition onto an oligounucleotide being used for primingallows synthesis to remain both efficient and inexpensive. At the sametime, they are sufficiently long that they are unique compared to thegenomic DNA they are being used with. Also, the phage RNA polymerasesthat recognize these promoters are usually single protein molecules thathave no requirement for other subunits or cofactors. Of special use inthis aspect of the present invention are phage promoter sequences thatare recognized by the T3, T7 and SP6 RNA polymerases. These enzymes arewell characterized and are commercially available from a number ofsources.

[0180] For efficient functionality, the promoters cited as examplesabove should be in double-stranded form. This may be carried out inseveral different ways. A potential sequence of events for one suchmethod is graphically depicted in FIG. 4. If the polymerase used forextension has strand displacement activity, the primer binding closestto the 3′ end of the 1^(st) strand (Primer A in FIG. 4) remains bound tothe template, but the other extended primers (Primer B and Primer C) arereleased from the template in single stranded form. Thus, a givenindividual template molecule may give rise to a plurality ofcomplementary copies by multiple priming/extension events with twogroups of products: essentially double-stranded molecules that comprisethe 1^(st) cNA strands bound to their complements and single-strandedmolecules derived from the displaced strands.

[0181] Although initially the displaced strands are in single-strandedform, the continued presence of other primers from either 1^(st) or2^(nd) strand synthesis could allow further binding/extension eventsthat convert the displaced single strands into double-stranded form.Alternatively, there may have been intermediary purification steps takento separate extended primers from non-extended primers. For example,separation may be useful to minimize or prevent the synthesis ofmolecules with promoters at each end. Such double-ended constructs maynot transcribe efficiently or may produce nucleic acids that hybridizewith each other rather than the target elements of the array. Therefore,the same primers that were used to initiate synthesis of the 1^(st) cNAstrand can be added to the mixture with the displaced 2^(nd) cNA strandsas well as whatever reagents may also be necessary to convert thedisplaced single-stranded DNA molecules into double-stranded products.Alternatively, random primers without promoters may be used for primingthe displaced 2^(nd) cNA strands. The synthesis of a complementary copyfor the displaced single strands also converts the promoter segment inthe 5′end of these molecules into double-stranded form.

[0182] On the other hand, the promoter in the extended primer thatremains bound to the original 1^(st) cNA strand template (Primer A inFIG. 4) needs different processes to render it into a functionallyefficient form. For instance, the single-stranded 3′ tail of the 1^(st)cNA strand could be digested by the 3′ to 5′ Exonuclease activity of T4DNA polymerase. Upon reaching the double stranded portion, the enzymecould then use its polymerase activity to extend the shortened 3′ end byusing the promoter segment of primer A as a template thereby generatinga double-stranded promoter. In another approach, oligonucleotides can beprovided that are complementary to the single-stranded promotersequences (FIG. 5a) or the primers used for 2^(nd) strand cNA synthesiscan be designed such that they are self-complementary and form stem loopstructures that generate double-stranded functional promoters (FIG. 5b).Lastly, the 2^(nd) cNA strands bound to the template can be denaturedand the same processes described above for converting the displaced2^(nd) cNA strands can be used to convert them into double-strandedform.

[0183] The creation of functional transcriptional units from theoriginal diverse nucleic acid analytes allows amplification by makingmultiple transcript copies from each cNA template. By inclusion of theRNA promoter sequence in primers that used the 1^(st) cNA strand as atemplate, all the resultant transcripts are also complementary to the1^(st) cNA strand. However, some target arrays that use definedoligonucleotide sequences as target elements have been designed for thepurpose of detecting labeled 1^(st) cDNA copies of mRNA rather thantheir complements. In such a case, the transcription products of theseries of reactions described above can be used as templates tosynthesize sequences equivalent to labeled 1^(st) cDNA copies by reversetranscription. As described previously, random or selected primers mayfind use for this purpose. This conversion step may offer otheradvantages as well since DNA is known to be more stable than RNA and hasrelatively less secondary structure compared to RNA.

[0184] RNA transcripts or cDNA copies of the RNA transcripts createdfrom the processes described above can either be labeled or unlabeled.When the polynucleotides are unlabeled, they can use UDTs for signalgeneration. As described previously, the original anlytes may haveinherent UDT sequences that may serve this function or the analytes maybe modified by the incorporation of non-inherent UDT sequences. On theother hand, the synthetic steps that are carried out in the series ofreactions above provide the opportunity to incorporate non-inherent UDTsduring either 1^(st) strand or 2^(nd) strand synthesis by primers withappropriate designs. For example, a primer design for 2^(nd) strandsynthesis can have the following structure:

[0185] 5′ promoter-UDT-hexamer-3′.

[0186] After binding the primer above to a 1^(st) cNA strand followed byextension, the transcripts could be generated with the structure:

[0187] 5′ UDT-hexamer-RNA sequence-3′.

[0188] Although the transcript shown above has a UDT at its 5′ end,other designs allow the transcripts to be synthesized with UDTs in their3′ ends. For instance, this can take place by either the sequence of theprimer binding site used for the initial 1^(st) strand synthesis beingcapable of acting as a UDT or by incorporation of a UDT into the primerthat is to be used for 1^(st) strand synthesis. As an example of bothmethods, a transcription unit can be synthesized from poly A RNA bypriming of the 1^(st) cNA strand with an oligonucleotide primer with thestructure:

[0189] 5′ UDT OligoT-3′

[0190] and priming of the 2^(nd) cDNA strand by an oligonucleotideprimer having the structure

[0191] 5′ promoter-hexamer-3′.

[0192] The double-stranded product of 1^(st) cNA and 2^(nd) cNA strandsynthesis reactions would then have the following structure:

[0193] 5′ promoter-hexamer-2^(nd) strand sequence-PolyA-UDT 3′

[0194] Transcription from this construct would generate RNA moleculesthat have the following structure:

[0195] 5′ hexamer-2^(nd) strand sequence-PolyA-UDT 3′

[0196] The product above can bind a UDE either through the an inherentUDT (the Poly A sequence) or through the artificially incorporated UDT.In addition, it should be recognized that the incorporation of UDTs forsignal generation can be coupled with incorporation of labelednucleotides if desired. Thereby, either by direct labeling or by thepresence of UDTs, this aspect of the present invention provides for thesynthesis of a library of detectable products that will reflect theinitial levels of the various nucleic acid analytes of a library.

[0197] The use of amplification utilizing RNA synthesis has beenpreviously described by Kwoh and Gingeras, (1989, Proc. Nat. Acad. Sci.USA 86; 1173-1177; incorporated herein by reference) but the purpose ofthat work was in diametric opposition to the present invention. In Kwohand Gingeras, primers with specific sequences were used to synthesizethe 2^(nd) cDNA strand in order to amplify a single defined discretesequence that was of interest. Thus there is no suggestion orrecognition of potential benefits of amplification of a diversepopulation of various nucleic acids.

[0198] In a patent application that was filed in the same year as thepublication by Kwoh and Gingeras, a method was described by van Gelderet al. (U.S. Pat. No. 5,716,785; incorporated herein by reference) forlinear amplification of a general population of RNA targets by includinga phage promoter into the primer used for the 1^(st) cDNA strand.Synthesis of the 2^(nd) strand were carried out either by nicking of theRNA template by RNase H or by hairpin formation at the end of the 1^(st)cDNA strands to provide self-priming events. Furthermore, the claims forthis patent and a related patent by the same inventors (U.S. Pat. No.5,891,636; incorporated herein by reference) specifically includes thephrase “without using an exogenous primer”. Thus, in these patents thereis firstly a requirement of inclusion of a promoter sequence into theprimers used for 1^(st) strand synthesis. Secondly there is noappreciation for the use of primers being added to catalyze the 2^(nd)strand synthesis. In fact, there is even a teaching away from thislatter concept. In addition, all of the foregoing methods synthesizeincomplete copies of the primary analytes as the completeness of thecopies made by RNase H are dependent upon the distance of the nick thatis closest to the 5′ end of the mRNA, only a minority will haverepresentation of the sequences closest to the 5′ end of the mRNA. Inaddition, there would never be representation of the end itself since itwould be used for retaining the RNA fragment/primer closest to the 5′end. Synthesis by means of hairpin formation also has intrinsicallyincomplete representation of the 5′ end sequences since nucleasedegradation of these sequences takes place during elimination of thehairpin. Also, there may be other losses since even nucleases that areconsidered to be single strand specific are more accuratelycharacterized as having a preference for single-strands since it is wellknown that there is also some level of activity with segments that arein double-stranded form.

[0199] The present invention is in contrast to previously cited art thatdid not use primers for 2^(nd) strand synthesis. These methods ofprevious art depended upon the presence of RNaseH to create a secondstrand or else required self-priming events by a foldback mechanism andsubsequent treatment with S1 nuclease or its equivalent. In the absenceof such a nuclease treatment, transcripts made from hairpin derivedconstructs would be self-complementary and thus incapable of appreciablehybridization to arrays. In contrast to this prior art, the presentinvention discloses various methods where exogenous primers are used tosynthesize the 2^(nd) strand. Also, in some aspects of the presentinvention, the methods used to synthesize the 2^(nd) strand includemeans that selectively retain information from the 5′ ends of analytes.In addition, the present invention describes the potential for thesynthesis of multiple transcription units from a single 1^(st) strandcNA template thereby providing an additional level of amplification.

[0200] It is another aspect of the present invention that the 1 st cNAstrands can be actively prevented from creating 2^(nd) cNA strandsthrough a fold-back mechanism by blocking the extendability of a 1^(st)cNA strand. One method of carrying this out is by the addition of adideoxynucleotide to the 3′ terminus of a 1^(st) cNA copy by terminaltransferase. Although this method would prevent a 1^(st) cNA strand fromparticipating in self-priming reactions, a blocked 1^(st) can strandwould retain its capability of being used as a template. In this aspectof the present invention, either the primer used for 1^(st) strand cNAsynthesis or 2^(nd) strand cNA synthesis can comprise an RNA promoter orother replication center.

[0201] Another aspect of the present invention discloses the addition orincorporation of artificial primer binding sites to carry out the novelprocesses described above. For instance, the translation of mRNA into acDNA copy also frequently includes the terminal addition of a fewnon-template directed nucleotides into the 3′ end of the 1^(st) cNAstrand by Reverse Transcriptase. In previous art, these added bases havebeen used as primer binding sites for cloning of full length cDNAmolecules. The addition of a few Cytosine nucleotides at the end of amolecule has been sufficient for the binding and extension of a primerthat has 3 Guanosine nucleotides at it 3′ end (user Manual for SMARTcDNA Technology, Clontech Laoboratories, Inc., Palo Alto, Calif.). Inthis system, aborted or stalled cDNA sequences that were incompletecopies of the original mRNA molecules would not be substrates for theaddition reaction by Reverse Transcriptase. This provided for a morecomplete representation of the 5′ sequences of the original mRNA in alibrary of cDNA clones.

[0202] The non-template derived addition of Cytosine nucleotides to the1^(st) cDNA strand has been previously used in the process of making atranscription library (Wang et al. 2000, Nature Biotechnology 18;457-459; incorporated herein by reference). However, this system wasbasically similar to the method described by van Gelder et al., (op.cit.) since a phage promoter was included in the primers used forsynthesis of the 1^(st) cDNA strand. As such, this arrangement has thelimitation that it has lost the selectivity for molecules that havecopied completely their mRNA templates. Primers that bind to interiorpoly C sequence and initiate extensions are as competent as bindings topoly C's at the end of cDNA (Matz et al., 1999) to synthesize 2^(nd)cDNA strands, thereby creating functional double stranded phagepromoters.

[0203] In contrast to van Gelder et al., and Wang et al., thisparticular aspect of the present invention provides a promoter in theprimer used for the 2^(nd) strand synthesis. Thus, the novel processesthat have been disclosed previously can be carried out by the use of aprimer for 2^(nd) strand synthesis that comprises oligo dG sequences attheir 3′ end for binding to the termini of 1^(st) cNA strands. In thisaspect of the present invention, priming events that derive from theterminal bindings and extensions will lead to double stranded promotersin molecules. As illustrated in Step (D) in FIG. 6, a primer with a T7promoter can bind to the terminus of the 1^(st) cNA strand. Extension ofthis primer can create a double stranded molecule where the 3′ end ofthe primer is extended using the cDNA as a template and the 3′ end ofthe cNA is extended using the primer sequences as a template. The netproduct of such extensions would be a double stranded transcriptionunit. On the other hand, Step (E) of FIG. 6 shows the binding of aprimer with a T7 promoter to an internal segment of the cNA with. Inthis case, although there can be extension from the 3′ end of the primerto create a partially double-stranded molecule, the 3′ end of the cNA isunable to use the primer as a template, thus leaving the promoter in anon-functional single-stranded form.

[0204] One advantage of the system described above is that thenon-template addition of nucleotides can be carried out by enzymes thatare already present in the reaction mixture. On the other hand, ifdesired, Terminal Transferase can be added to increase control over thereaction and improve efficiency. When poly A, T or U sequences arealready present in either RNA, DNA or cNA copies, it is preferred thatthe Terminal transferase use dGTP or dCTP. Primers for 2^(nd) strandsynthesis can then be designed whose sequences comprise a promoter and a3′ segment complementary to the sequences added by the TerminalTransferase addition step. The steps of this process are shown in FIG.7, where subsequent extensions to create a double stranded promoter canbe carried out as previously described for FIG. 6. Also, since thedirected addition of nucleotides takes place only where there is eithera double stranded end or a free 3′ end, only cDNA molecules that havebeen completely extended to the ends of the analyte templates will besuitable substrates for terminal addition.

[0205] Since these additions can be longer than those derived fromnon-template additions by Reverse Transcriptase, the primers used for2^(nd) strand synthesis can have longer corresponding homopolymericsegments thereby allowing higher temperatures for binding and extension.This heightened stringency should decrease the frequency of primingevents with internal sequences in the 1^(st) cNA template strand andprovide higher representation of sequences from the 5′ end of theoriginal analytes. Therefore, when terminal transferase is used togenerate a primer binding site for 2^(nd) strand synthesis, the promotercan be in either the 1^(st) strand or the 2^(nd) strand. The step ofterminal transferase addition to the 1^(st) cNA can be carried out whileit is still bound to its template as described above, or it can becarried out after destruction of the template or separation of thetemplate from the 1^(st) cNA strand. This method should continue toenjoy 2^(nd) strand synthesis that is preferentially initiated byprimers binding and being extended from the 3′ termini of 1^(st) cNAstrands. As described previously, UDTs, as well as labeled or unlabelednucleotides can all be utilized in carrying out this aspect of thepresent invention. Also, it is contemplated that higher yields of endproducts can be achieved by repetitions of one or more steps of thevarious process that are disclosed herein.

[0206] Other means that preferentially carry out priming events at the3′ ends of 1^(st) strand cNA's may also find use in the presentinvention. For instance, a cDNA copy that is a complete copy of its RNAtemplate is a substrate for blunt end ligation by T4 DNA ligase with adouble-stranded oligonucleotide. The sequence of the oligonucleotideligated to the 3′ end of the 1^(st) cNA strand can be chosen by the userand can function as a primer binding site for making a 2^(nd) cNAstrand. Similarly a 3′ single-stranded tail in the 1^(st) cNA strand isa substrate for ligation of a single-stranded DNA oligonucleotide by T4RNA ligase (Edwards et al., 1991 Nucleic Acids Research 19; 5227-5232;incorporated herein by reference). Lastly, a double-strandedoligonucleotide with a 3′ single-stranded tail can be joined to a 1^(st)strand cNA through “sticky end” ligation by T4 DNA ligase when the1^(st) cNA and oligonucleotide tails are complementary. As describedpreviously, these cNA tails can be derived from non-template additionsby Reverse Transcriptase or by Terminal transferase. Illustrativeexamples of these processes are given in FIG. 8. Since all of theseprocesses are dependent upon preferential binding of primers to the 3′ends 1^(st) strand can molecules, the promoter can be in either the1^(st) or 2^(nd) cDNA strand.

[0207] In another embodiment of the present invention, a 1^(st) strandcNA strand is fragmented by physical, chemical or enzymatic means.Examples of enzymatic means can include but not be limited torestriction enzymes such as Hha I, Hin P1 I and Mnl I, DNases such asDNase I and nucleases such as S1 nuclease and Mung Bean Nuclease. Thesefragments can be used as templates for synthesis of a 2^(nd) strand byany of the methods described previously. For example, hybridization andextension of random primers with T7 promoters can be used with the cNAstrand fragments as templates in processes similar to those shown inFIGS. 4 and 5. Or if preferred, the homopolymeric addition or ligationsteps described above can be carried out to provide specific primerbinding sites. FIG. 8 is an illustration of this process using thehomopolymeric method. Breaking down the 1^(st) strand copy into smallersegments followed by incorporation of a primer during 2^(nd) strandsynthesis would provide smaller transcription units. This may beadvantageous when using modified nucleotides for signal generation. Forinstance, when there are long stretches in the template strand that arecomplementary to the labeled nucleotide, the modification to thenucleotide may cause a blockage in downstream transcription or loss ofprocessivity and result in under-representation of those sequences. Inthis particular aspect of the present invention, the partition of copiesof analyte sequences into smaller individual transcription units allowseach of the units to direct RNA synthesis independently thereby creatinga more complete representation of the library of various nucleic acidsequences.

[0208] In another embodiment of the present invention, the novel methodsdisclosed for synthesis of a library are combined with capture methodsto provide more efficient synthesis as well as flexibility in changingsalts, buffers, enzymes and other components during multistep processes.The present invention discloses the use of a 1^(st) strand primer thatis bound to a solid matrix such as a bead followed by the processesdescribed above. For example, the 3′ end of Oligo T sequences bound to asolid matrix can be extended using polyA mRNA as a template. Inaccordance with the methods of the present invention, this 1^(st) cNAstrand is thereupon used as a template for the 2^(nd) cNA strand. Whencarrying out this aspect of the present invention, a replicative centersuch as an RNA promoter sequence can be introduced into either the1^(st) or 2^(nd) strand depending upon the particular method used. Forinstance, random primers with promoters in their 5′ ends can bind to theextended 1^(st) cDNA strands to create 2^(nd) strands that have apromoter incorporated into them. This process is depicted in FIG. 10.

[0209] The single-stranded promoter on the 5′ ends of the 2^(nd) cDNAstrands can be converted into double-stranded form by any of the methodsdescribed previously. For instance, the primer/template complex thatremains bound to the bead in FIG. 10 can be treated with T4 DNApolymerase, hybridized with an oligonucleotide complementary to thepromoter segment or the primer can be designed with self complementaryregions. The latter two methods were previously discussed with referenceto FIG. 5. With regard to the displaced 2^(nd) cDNA strands in FIG. 10,the presence of unextended oligo-T tails on the matrix material canprovide further binding/extension events since the displaced strandscarry poly A sequences on their 3′ ends. However, if preferred, moreoligo-T can be added whether associated with beads or free in solution.Extension of the oligo-T should ultimately result in conversion of thesingle-stranded promoters of the displaced 2^(nd) cDNA strands intofunctional double-stranded forms.

[0210] Another method that can be used in the present invention is torepeat one or more of the steps that have been described in the presentinvention. For instance, after using a library of analytes to synthesize1^(st) can copies attached to a matrix, the anlytes can be separatedfrom the 1^(st) cNA copies and used to create another pool of 1^(st) cNAcopies. Similarly, after synthesis of 2^(nd) can strands, the library of2^(nd) cNA strands can be separated from the 1^(st) can strands fixed tothe matrix. All 2^(nd) cNA strands that have copied the 5′ ends of the1^(st) cNA strands will have regenerated the sites that were initiallyused to bind to the primers linked to the beads. If desired, the 2^(nd)strands can be rebound to the same beads. Since there are likely to bean enormous number of poly T primers on the beads compared to the numberof templates used for 1^(st) cNA synthesis, the majority of primers onthe matrix remain unextended and can be used for new priming events.Thus, complete copying of these rebound 2^(nd) can strands should allowgeneration of double-strand promoters at the ends of these moleculeswithout a necessity for the use of T4 to do “trimming”. If desired the1^(st) cNA strands that are attached to the matrix can be used togenerate another pool of 2^(nd) cNA strands. The pool or pools of 2^(nd)can strands can then be added to fresh beads with primers complementaryto their 3 ′ ends. Again, the extension of the primers attached to thematrix will convert all of the 2^(nd) can strands into double-strandedform including the promoter sequences that were at their 5′ ends.Lastly, after a transcription reaction is carried out, the reactionproducts can be removed and the nucleic acid on the matrix can be usedfor more transcription reactions thereby accumulating more transcriptionproducts.

[0211] Although the example above describes priming of an analyte with apoly A segment by an oligo T primer attached to a matrix, thee primerscan also be prepared with one or more discrete bases at their 3′ ends.As described previously, these primers can be used as a group thatrepresents all the possible variations or they can be used individuallydepending upon whether general amplification or separation intosubclasses was desired. The poly A sequence used above is understood toonly be an illustrative example. As described previously, the sequencesin analytes used for binding of 1^(st) strand primers can be derivedfrom inherent sequences or they may be noninherent sequences in analytesthat have been artificially introduced by any of the means that havebeen described previously. This particular embodiment of the presentinvention can utilize any of these primer binding sites by appropriatedesign of the primer sequence bound to the matrix.

[0212] In the present invention, the primer sequences for 1^(st) strandsynthesis can be either directly or indirectly attached to a matrix.Methods for direct attachment of oligonucleotides to matrixes are wellknown in the art. In addition, beads with covalently attached extendablepoly T segments are commercially available from a number of sources.Methods for indirect attachment are also well known in the art. Forinstance FIG. 11 depicts a sandwich method where a primer has twosegments, one of which is complementary to a capture segment attached tothe matrix and the other is complementary to the poly A segment of thetarget RNA. The two segments of the primer may form a continuousnucleotide sequence or there may be a disjunction between the twosegments. Hybridization of the two segments of the primer and thecomplementary sequences on the matrix and the binding site of theanalyte can take place simultaneously or they can be carried out in astep-wise fashion. For instance, hybridization of target RNA to thecapture element can be carried out in solution followed by capure to thematrix. It is preferred that the segment that is bound to the matrix berendered incapable of extension. One way this blockage can be carriedout is by the use of the 3′ end as the attachment point to the matrix asdepicted in FIG. 11. Binding and extension events can take place asdescribed previously for FIG. 10 to synthesize 1^(st) and 2^(nd) cDNAcopies of the original poly A mRNA. Conversion of the promoter sequencesinto double-stranded form can also take place as described above.Transcription can take place either while the transcription units areattached to the matrix or if desired separation from the matrix can takeplace in a step subsequent to the transcription.

[0213] Incorporation of an RNA promoter during 1^(st) strand synthesisresults in transcripts that comprise sequences that are complementary tosequences in the original analytes. Incorporation of an RNA promoterinto the 2^(nd) strand synthesis results in the production oftranscripts that comprise sequences that are identical to sequences inthe original analytes. As described previously, these can easily beconverted into complementary cDNA copies if desired.

[0214] It is a further subject of the present invention thattranscription units can be synthesized without incorporating a promotersequence into either the 1^(st) cNA (as described by Eberwine et al.,op. cit.) or the 2^(nd) cNA strand (as described in previous embodimentsof the present invention). As shown in step D of FIG. 12, when usingextended 1^(st) cNA strands as templates for synthesis of the 2^(nd) cNAstrands, a duplicate of the original primer binding sequence issynthesized. Thus, in FIG. 12 a polyA segment is created at the 5′ endsfor both displaced 2^(nd) cNA strands and for 2^(nd) cNA strands thatremain bound to the beads. After removing these 2^(nd) cNA strands,oligonucleotide primers comprising an RNA promoter and oligo-T sequencescan be hybridized to the 2^(nd) cNA strands. The primers may be attachedto a matrix or they may be free in solution. Provision of DNAPolymerase, nucleotides and appropriate cofactors can allow extension ofboth the 3′ ends of the promoter/primers as well as the 3′ ends of thecDNA copies thereby creating functional transcriptional units as shownin step F of FIG. 12. Transcription from these DNA molecules will resultin products that comprise sequences that are complementary to sequencesin the original analytes In previous art the most common use of oligo-Tthat is attached to a matrix such as cellulose or beads has been for thepurpose of a selective isolation of polyA mRNA followed by a releasestep prior to synthesis of a library. In one instance, a special oligo Tprimer joined to a T7 promoter was extended using RNA as template tocreate a library (Eberwine op.cit.). However, this system put thepromoter in close proximity to the capture bead, potentially decreasingits ability to be converted into double-stranded form and/or for it tofunction as a promoter. Also, synthesis of the 2^(nd) strand by randompriming does not prevent hairpin self-priming. In the absence of anuclease step, transcription units would direct synthesis ofself-complementary RNAs from hairpin template sequences that would beincapable of hybridizing to target arrays. use of the templates for thisnon-productive synthesis may cause an inefficiency in the amount ofeffective labeled transcripts

[0215] A particular benefit of the use of promoters in primers used for2^(nd) cNA synthesi present invention is that although 1^(st) cNAstrands can be synthesized under conditions that have the potential forself-priming events i.e. creating 2^(nd) cDNA strands by a fold-backmechanism, the absence of a promoter in 1^(st) cDNA; strand wouldprevent these constructs from being transcriptionally active. Thus, only2^(nd) cDNA strands that are derived from priming events byoligonucleotides with promoter sequences are functional fortranscription. This in contrast to the system previously described byEberwine (op. cit.). Contrariwise, methods have also been described inthe present invention that allow the use of a promoter in the 1^(st)strand by either preventing extension of a 1^(st) cNA strand or byfacilitating 2^(nd) strand synthesis from priming events at the ends of1^(st) strand templates.

[0216] It is another object of the present invention to provide a methodfor comparative analysis that requires only a single RNA population tobe labeled. This particular aspect takes advantage of competitivebinding by an unlabeled population of RNA. Synthesis of this materialcan take place by any of the means described in the foregoing work. Theparticular sequences can be homologous to sequences that are present onthe arrays or they may be homologous to sequences that are present inthe labeled material. By comparison of hybridization of the labeledmaterial in the presence or absence of competitor, relative levels ofincreased or decreased mRNA synthesis can be established relative to thecompetitor, ie. differential competition. Adjustments can be made in therelative amounts of unlabeled material being used or the housekeepinggenes that are present as controls can allow for normalization values.This method provides the advantage that multiple sequential or parallelhybridizations can be carried out and compared with a single commonlabeled control population of RNA.

[0217] The various steps of the present invention can be carried outsequentially by adding various reagents and incubation steps asrequired. On the other hand, the series of steps can be segregated byintroducing additional steps that either remove or inactivate componentsof the reaction or where a desired product is separated from a reactionmixture. An example of the former can be heat inactivation of ReverseTranscriptase. An example of the latter can be isolation of RNA/DNAhybrids by selective matrices. These additional steps can be carried outto either improve the efficiency of subsequent steps or for the purposeof preventing undesirable side reactions.

[0218] Although the previous examples have disclosed the utility of aphage promoter in carrying out various aspects of the present invention,a production center is able to operate by other means as well. Forinstance, various means of introducing UDTs that serve as primer bindingsites have been previously described in the context of synthesis of2^(nd) copy strands followed by RNA transcription. These primer bindingsites can in themselves serve as production centers for multiple copiesof various nucleic acids under isothermal conditions.

[0219] For instance the use of primers that are designed to createtarget-dependent stem-loop structures has previously been disclosed inRabbani et al., U.S. patent application Ser. No. 09/104,067, filed onJun. 24, 1998 (Novel Processes for Amplifying Nucleic Acid,Post-Termination Labeling Process for Nucleic Acid Sequencing andProducing Nucleic Acid Having Decreased Thermodynamic Stability; forspecific isothermal amplification of selected sequences. The content ofthe aforementioned Ser. No. 09/104,067 is hereby incorporated byreference. In the present invention, UDTs can be added to the variousnucleic acids of a library to carry out the amplification disclosed inRabbani et al., U.S. patent application Ser. No. 09/104,067, cited supraand incorporated herein by reference. FIG. 13 is a depiction of a seriesof reactions that could be used to carry this out. For instance, a UDTcan be ligated to a library of poly A mRNA where the UDT comprises twosegments (termed X and Y in this Figure). In the next step, a primer(Primer 1) that comprises two segments, a poly T sequence at the 3′ endand a segment termed Z at the 5′ end is hybridized to the poly Asequences at the 3′ end of the mRNA and extended by reversetranscription to make a 1^(st) cNA copy (Steps C and D of FIG. 13) thatcontains the sequnces X′ and Y′ at the 3′ end. Removal of the originaltemplate makes the X′ segment at the 3′ end of the 1^(st) cNA copyavailable for hybridization. A second primer (Primer 2) that has twosegments, segment X at the 3′ end and segment Y′ at the 5 ′ end can beannealed and extended to make a 2^(nd) copy (Steps D and E) of FIG. 12.The presence of Primer 2 should also allow a further extension of the1^(st) cNA copy such that a double stranded segment is formed where theY and Y′ segments are capable of self-hybridizing and thereby creating astem-loop structure with the X and X′ segments in the loop portions asdescribed in Rabbani et al., U.S. patent application Ser. No.09/104,067, cited supra and incorporated herein by reference. Creationof a stem loop at the other end can be carried out by annealing a thirdprimer (Primer 3) which comprises two segments, segment Z at the 3′ endand a Poly A segment at the 5′ end using a 2^(nd) cNA copy as atemplate. The availablity of 2^(nd) cNA copies as templates can bederived from multiple priming events by Primer 2 at the other end (asdescribed in Rabbani et al., U.S. patent application Ser. No.09/104,067, cited supra and incorporated herein by reference, or bydenaturation of the 1^(st) and 2^(nd) strands from each other. Extensionof Primer 3 creates a structure that has the Poly T and Poly A segmentsforming a stem and the Z and Z′ segments forming the loops. Furtherbinding and extension reactions under isothermal conditions can proceedas described previously for unique targets. It should be noted that theparticular sequences used for X, Y and Z are arbitrary and can be chosenby the user. For instance, if the Z segment of Primer 1 used in step Cof FIG. 13 was designed with X and Y sequences at the 5′ end, the unitlength amplicon would have X′ and Y segments at the 3′ end of eachstrand. As such, amplification could be carried out using only Primer 2.

[0220] Another example of the use of non-inherent UDTs being used asprimer binding sites for isothermal amplification is shown in FIG. 14for use with the Strand Displacement Amplification system described byWalker et al., in U.S. Pat. No. 5,270,184 herein incorporated byreference. In this particular example, Incorporation of segment X takesplace by two different methods. In step B of FIG. 14, segment X isintroduced by ligation to an analyte of the library. In step C segment Xis attached to a poly T primer and becomes incorporated by strandextension. The presence of the X segment at the 5′ end of each end ofthe amplicon unit allows primer binding by a single Strand Displacementprimer. Methods for the designs of primers with appropriate sequences attheir 5′ ends have been described by Walker et al., (op. cit.). Withregard to the particular enzyme being used as part of the SDA system,the presence of a particular restriction site between primer bindingsites may limit the ability of some sequences to be amplified in areaction designed for general amplification of a library. This may beovercome by choosing relatively uncommon sequences or carrying outparallel reaction with different enzymes.

[0221] It should be pointed out that in the examples shown in FIGS. 13and 14, the presence of primer binding sites at each end allowsexponential amplification. However, these processes can be changed tolinear amplification by designing amplicons that have binding sites forisothermal amplification at only one end of the amplicon.

[0222] Incorporation of a primer binding site that can be used forisothermal production of multiple copies can take place by any of thesteps described previously that used a promoter in the example. Forinstance, FIGS. 13 and 14 show addition of an isothermal binding sitedirectly to an analyte and also show incorporation of an isothermalbinding site during synthesis of a first copy. FIG. 15 shows a similarsituation, but in this example segment X is incorporated during 1^(st)cNA synthesis, segment Q is added after first strand synthesis andsegment Z is added during 2^(nd) cNA strand synthesis. As describedpreviously, one or more of these segment can comprise primer bindingsites for isothermal synthesis. It should also be pointed out that inFIGS. 13 through 15 both inherent and non-inherent UDTs were used aspart of the examples.

[0223] In another aspect of the present invention, UDTs are used asprimer binding sites for amplification on an array. In this particularaspect, each locus on an array comprises two sets of primers. The firstset of a locus comprises Selective Primer Elements (SPE's) that arespecific for a particular analyte. The second set of a locus comprisesUniversal Primer Elements (UPE's) that are identical or complementary tosequences in UDT elements. As described previously, UDTs can be derivedfrom naturally occurring sequences or they may be artificiallyincorporated. The SPE”s at a locus would be able to bind to thecomplementary sequences in the nucleic acids of a library, therebybinding discrete species of nucleic acids to that particular locus ofthe array. The use of appropriate conditions, reagents and enzymes wouldallow an extension of an SPE using the bound nucleic acid as a template.

[0224] As an example of this aspect of the present invention, FIG. 16depicts an array with three different loci termed Locus P, Locus Q andLocus R. At each of the loci, there is a set of SPE's bound to the arraythat are complementary to a particular sequence in cDNA copies made fromone of three species of poly A mRNA termed P, Q and R respectively. Inaddition, each locus of the array in FIG. 16 has a set of UPE's thatcomprises poly T sequences. Synthesis of a cDNA copy of each of the mRNAtemplates by Poly T priming of their polyA tails creates cDNA P, cDNA Qand cDNA R respectively. Binding of the 1^(st) cDNA strand of an analyteto an SPE should be selective for each species at a particular locus. Onthe other hand, there should be little or no binding of the cDNA copiesto the universal Poly T sequences in the UPE's of the array of FIG. 16.The addition of enzymes and reagents for extension should generate2^(nd) cDNA copies of P, Q and R at the LP, LQ and LR sites on the arrayby extension of SPE's using the bound cDNA as templates. Each of these2^(nd) cDNA copies would comprise unique sequences complementary to the1^(st) cDNA strand templates. However, in addition to these uniquesequences, the 2^(nd) strand copies would include a common poly Asequence at their 3′ ends. At this stage it may be preferable to removeunhybridized analytes as well as templates used for 2^(nd) strandsynthesis. This is most easily carried out by heat denaturation followedby washing steps. The product at this stage is an array that hasextended and un-extended SPE's at each locus where the number ofextended SPE's should be in proportion to the amount of the originalcorresponding analytes. The extended SPE's can now serve as templateswhen an unextended poly T UPE is in sufficient proximity. The design andplacement of pairs of unique primers for solid phase amplification hasbeen previously described in detail in U.S. Pat. No. 5,641,658, herebyincorporated by reference. Methods for synthesis of arrays with twodifferent sequences at each locus has also been described by Gentalenand Chee, 1999 (Nucl. Acids Res. 27; 1485-1491) incorporated byreference. The same primer design rules may also be applied to thepresent invention that uses non-unique primers. Extension of a UPE witha nearby extended SPE as a template creates a new template that can inturn be used as a template for a nearby unextended SPE. This process canproceed through a series of binding and extension steps thatalternatively using SPE's and UPE's to accumulate nucleic acids that arederived from target nucleic acids homologous to the sequences in the SPEat each locus. An illustration of these steps is given in FIGS. 16through 19.

[0225] Methods for the design and synthesis of arrays for solid phaseamplification have been described in U.S. Pat. No. 5,641,658 and Weslinet al., 2000, (Nature Biotechnology 18; 199-204; both documentsincorporated herein by reference) for utilization of totally unique setsof primers. Methods of assaying the extent of synthesis are alsodescribed in these references. For example, labeled precursors can beincluded in the reaction to synthesize a labeled amplification product.Alternatively, normal precursors can be used with signal generationprovided by intercalating dyes binding to amplification products.

[0226] This invention provides unique compositions and processes forsolid phase amplification. Among such compositions is one that comprisesan array of solid surfaces comprising discrete areas, wherein at leasttwo of the discrete areas each comprises a first set of nucleic acidprimers; and a second set of nucleic acid primers; wherein thenucleotide sequences in the first set of nucleic acid primers aredifferent from the nucleotide sequences in the second set of nucleicacid primers; wherein the nucleotide sequences of a first set of nucleicacid primers of a first discrete area and the nucleotide sequences of afirst set of nucleic acid primers of a second discrete area differ fromeach other by at least one base; and wherein the nucleotide sequences ofthe second set of nucleic acid primers of a first discrete area and thenucleotide sequences of the second set of nucleic acid primers of asecond discrete area are substantially the same or identical. Previousdescriptions for any of the above-recited elements have been givenelsewhere in this disclosure, and resort may be made to thosedescriptions in connection with this process.

[0227] A related composition of this invention is one comprising anarray of solid surfaces comprising a plurality of discrete areas;wherein at least two of the discrete areas each comprises a first set ofnucleic acid primers; and a second set of nucleic acid primers; whereinthe nucleotide sequences in the first set of nucleic acid primers aredifferent from the nucleotide sequences in the second set of nucleicacid primers; wherein the nucleotide sequences of a first set of nucleicacid primers of a first discrete area and the nucleotide sequences of afirst set of nucleic acid primers of a second discrete area differsubstantially from each other; and wherein the nucleotide sequences ofthe second set of nucleic acid primers of a first discrete area and thenucleotide sequences of the second set of nucleic acid primers of asecond discrete area are substantially the same or identical. See thisdisclosure above and below for a description of any of the elements inthis process.

[0228] Related to the last-mentioned compositions are processes forproducing two or more copies of nucleic acids of interest in a librarycomprising the steps of a) providing (i) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of thediscrete areas each comprises: (1) a first set of nucleic acid primers;and (2) a second set of nucleic acid primers; wherein the nucleotidesequences in the first set of nucleic acid primers are different fromthe nucleotide sequences in the second set of nucleic acid primers;wherein the nucleotide sequences of a first set of nucleic acid primersof a first discrete area and the nucleotide sequences of a first set ofnucleic acid primers of a second discrete area differ from each other byat least one base; and wherein the nucleotide sequences of the secondset of nucleic acid primers of a first discrete area and the nucleotidesequences of the second set of nucleic acid primers of a second discretearea are substantially the same or identical; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacids of interest; b) contacting a primer of the first set with acomplementary sequence in the nucleic acid of interest; c) extending theprimer in the first set using the nucleic acid of interest as a templateto generate an extended first primer; d) contacting a primer in thesecond set with a complementary sequence in the extended first primer;e) extending the primer in the second set using the extended firstprimer as a template to generate an extended second primer; f)contacting a primer in the first set with a complementary sequence inthe extended second primer; g) extending the primer in the first setusing the extended second primer as a template to generate an extendedfirst primer; and h) repeating steps d) through g) above one or moretimes. Elements above are described elsewhere herein.

[0229] Another related process useful for detecting or quantifying morethan one nucleic acid of interest in a library comprises the steps of a)providing (i) an array of solid surfaces comprising a plurality ofdiscrete areas; wherein at least two of such discrete areas eachcomprises: (1) a first set of nucleic acid primers; and (2) a second setof nucleic acid primers; wherein the nucleotide sequences in the firstset of nucleic acid primers are different from the nucleotide sequencesin the second set of nucleic acid primers; wherein the nucleotidesequences of a first set of nucleic acid primers of a first discretearea and the nucleotide sequences of a first set of nucleic acid primersof a second discrete area differ from each other by at least one base;and wherein the nucleotide sequences of the second set of nucleic acidprimers of a first discrete area and the nucleotide sequences of thesecond set of nucleic acid primers of a second discrete area aresubstantially the same or identical; (ii) a library of nucleic acidanalytes which may contain the nucleic acids of interest; (iii)polymerizing means for synthesizing nucleic acid copies of the nucleicacids of interest; and (iv) non-radioactive signal generating meanscapable of being attached to or incorporated into nucleic acids; b)contacting a primer of the first set with a complementary sequence inthe nucleic acid of interest; c) extending the primer in the first setusing the nucleic acid of interest as a template to generate an extendedfirst primer; d) contacting a primer in the second set with acomplementary sequence in the extended first primer; e) extending theprimer in the second set using the extended first primer as a templateto generate an extended second primer; f) contacting a primer in thefirst set with a complementary sequence in the extended second primer;g) extending the primer in the first set using the extended secondprimer as a template to generate an extended first primer; h) repeatingsteps d) through g) above one or more times; and i) detecting orquantifying by means of the non-radioactive signal generating meansattached to or incorporated into any of the extended primers in stepsc), e), g), and h). Elements above are described elsewhere herein.

[0230] For many uses, the UPE's will be present on the array duringhybridization of the analyte to complementary SPE's. However, there maybe circumstances where the presence of UPE's in this step may bedeleterious. For example, binding of the diverse nucleic acids of alibrary should preferably take place only through the action of theSPE's on the array. In contrast to the example given above, there may becases where due either to the nature of the library or the choice of UPEsequences, hybridization can take place between the library and theUPE's of an array. This event could result in a loss of efficiency inthe reaction by binding of target nucleic acids to inappropriate areasof the array. For instance, the SPE's at a particular locus would beunable to use complementary nucleic acid targets as a template if thesetargets are inappropriately bound to another physical location throughbinding of UPE's,. Furthermore, UPE's would be rendered non-functionalby being extended and synthesizing nucleic acid copies that lackcomplementary to the SPE's at that particular locus.

[0231] Accordingly, it is a subject of the present invention that UPE'smay be either non-functional or absent during the initial hybridizationof a library to the SPE's in the array. In one method of carrying thisout, advantage is taken of the universal nature of the UPE's. Althougheach particular species of SPE is relegated to a specific area of thearray, the UPE's are intended to be present in multiple areas of thearray. As such, an array can be synthesized where each locus comprises aset of SPE's and a set of chemically activated sites that are compatiblewith reactive groups on UPE's. After the initial hybridization ofnucleic acid targets to their appropriate SPE's, the UPE's withappropriate groups can be added universally to the array by asimultaneous attachment to all of the active sites on the array. Anexample of compatible modifications that could be used in this aspect ofthe present invention could be arrays that have maleimide groups at eachlocus and UPE's that have amine groups attached to their 5′ ends.

[0232] An alternative approach is for synthesize the array with UPE'sthat have been modified such that they are temporarily unable tofunction. For example, the UPE's could be synthesized with 3′ PO₄ groupsthereby blocking any potential extension reactions. After hybridizationof nucleic acids to the various SPE's of the array followed by extensionof SPE's, the nucleic acids used as templates could be removed from thereaction. After this step, the 3′ end of the UPE's could be renderedfunctional by removal of the 3′ PO₄ groups by treatment with reagentssuch as bacteriophage polynucleotide kinase or alkaline phosphatase.Thereafter, successive reactions can take place as described previously.

[0233] An alternative approach would be the use of hybridizationproperties of nucleic acids. For example, the Tm of hybridizationbetween nucleic acids is a function of their length and basecomposition. Therefore, the SPE's and UPE's can be designed with Tm'sthat are sufficiently different that salt or temperature conditions canbe used that selectively allow hybridization of the nucleic acids in thesample to SPE's. The salt and temperature conditions can be alteredlater to allow hybridization to the UPE's on the array and carry out theappropriate series of reactions.

[0234] Another example would be the use of competitive hybridization.Nucleic acids or their analogues can be added that are homologous to theUPE's. By either pre-hybridization or by including a high excess of suchcompetitors, the UPE's should all be occupied with the competitornucleic acids thereby allowing binding of the nucleic acids of thelibrary to SPE's only. Furthermore, the competitors can be synthesizedin such a way that even though they are bound to the UPE's they areunable to serve as templates for extension of the UPEs. Examples ofmeans that can be used for this purpose can include but not be limitedto peptide nucleic acids and oligonucleotides with multiple abasicsites. After extension of the SPE's, both the templates used forextension of SPE's and the competitor oligonucleotides bound to theUPE's can be removed concurrently rendering both the extended SPE's andUPE's available for binding to each other.

[0235] The poly A RNA in the example shown in FIGS. 16-19 made use of aninherent UDE in eucaryotic mRNA. As described previously, UDEs can alsobe added artificially either by polymerization or ligation. Forinstance, a selected arbitrary sequence can be added to the 5′ ends of alibrary of RNA analytes by the action of T4 RNA ligase. An array couldthen be used that has SPE's for unique RNA sequences and UPE's with thesame sequences as the ligated segment. After localization of the variousspecies of RNA to their appropriate location on an array, an enzymeappropriate for reverse transcription can be added as well as theappropriate buffers and reagents to extend the SPE's therebysynthesizing 1^(st) strand cDNA copies linked to the array. Removal ofthe RNA template would then allow the complement of the UPE in the cDNAcopy to bind to a nearby UPE on the array followed by a set of reactionsas described previously. Since the choice of sequences for artificiallyadded UPE's is of arbitrary nature, this aspect of the present inventioncan be applied to a simultaneous assay of different pools of analytes byadding different discrete UPE sequences to each library. In contrast tothis, the prior art cited above makes no provision for distinguishingbetween collection of analytes from different sources that have the samesequences. An illustration of an array that could be used for thispurpose is given in FIG. 20 where two libraries are being compared. Onelibrary has been prepared by joining sequences for UPE 1 to the nucleicacids and a second library has been prepared that has sequences for UPE2 joined to the nucleic acids. It should be noted that in FIG. 20, Locus1 of the array has the same SPE's as Locus 9 but they differ in theidentity of the UPE where UPE 1 is at Locus 1 and UPE 2 is at locus 9.This is also true for Locus 2 compared to Locus 10 and so on. Thus,binding of the same sequence can take place at either Locus 1 or Locus9, but the extent of amplification that will take place at each locuswill be dependent upon the amount of bound material that contains theappropriate UPE sequence.

[0236] In addition, although the examples above have used RNA or cDNAcopies as libraries for this aspect of the present invention, it hasbeen previously disclosed that DNA may also be the initial analyte. Asan example of this aspect of the present invention, DNA can be digestedwith a restriction enzyme to create a library of fragments. Adouble-stranded UDE can then be ligated to these fragments by the actionof T4 DNA ligase. The ligated products can then be denatured andhybridized to an array of SPE's. For example, to investigate potentialSNP's at a site “X” on a target nucleic acid, sets of SPE's can bedesigned that differ by a single nucleotide at their 3′ ends. Thesubsequent efficiency of extensions would then be dependent on how wellthe nucleotide at site “X” of the target template matched the 3′ base ofthe SPE. As an internal control, a set of SPE's can be designed thatwill utilize each strand at site “X”=0 thereby duplicating theinformation. This process is illustrated in FIG. 21. In this particularexample, it is preferred that binding between the nucleic acid and theUPE on the array be prevented since the ligated fragments will havesequences complementary to the UPE's. Examples of means that can be usedto carry this out have been described previously whereby UPE's areabsent or non-functional during hybridization of the nucleic acids tothe SPE's. On the other hands, the nucleic acids that are being analyzedcan be treated such that sequences that are complementary to UPE's areremoved. For instance, after the ligation step described above, nucleicacids can be treated with a 3′ to 5′ double-stranded Exonuclease. Thisshould selectively remove sequences complementary to the UPE's whileretaining sequences that are identical to sequences in the UPE's.Regeneration of the sequence complementary to the UPE should then takeplace only after extension of an SPE. Also as disclosed above, the useof artificial addition of UPE sequences allows the simultaneous analysisof different pools by a selective choice of different UPE sequences foreach pool.

[0237] It is a further intent of the present invention that rather thanchoosing specific sequences derived from prior sequence information, ageneral array can be made that offers complete representation of allpossible sequences. For instance, a library of SPE's that are 4 bases inlength with permutations of all 4 variable bases would comprise 4×4×4×4distinct sequences, i.e. a total of 256 permutations. With a complexityof all potential octamer oligonucleotides with the four variable bases,there would be a total of 256×256 for a total of 65,536 permutations. Inprior art, an array covering all the possible amplification productswould require two unique primers for each individual amplification.Thusly, there would be a requirement for a total of 65,536×65,536 for atotal of 4.3×10⁹ permutations for pairs of unique octamer primers on thearray. Such high numbers may be too expensive or too complex to havepractical application. On the other hand, the present inventionovercomes this limitation by virtue of the use of UPE's. Accordingly,only the SPE's need to encompass all the possible octamer sequenceswhich results in a requirement for a total of 65,536 differentsequences, a number that is easily within the ability of currenttechnology. The number of different nucleic acid that will be amplifiedat each locus will depend upon the complexity of the library of nucleicacids applied as templates as well as the conditions used for carryingout amplification. The degree of complexity of the array can also bealtered by increasing or decreasing the number of nucleotides comprisingthe SPE's. Conversely, it has previously been pointed out that a degreeof differentiation can be achieved by adding one or more discrete basesto the UPE. For example, the use of a single variable nucleotide at theend of a polyT UPE would decrease the complexity of the analytes in alibrary that could be amplified since on average, only one out of threeof the various diverse nucleic acid analytes bound to SPE's would beable to carry out strand extension. On the other hand, the inclusion ofall 3 sets of UPE's that each carries one of the 3 potential bases incombination with complete representation of octamer SPE's would increasethe complexity of arrays from 65,536 sequences to a total of 1.97×10⁵(3×65,536) permutations. By using variable nucleotides in the last twonucleotides at the 3′ end of the UPE on an array with SPE octamers, thecomplexity would be 8.0×10⁵ (12×65,536) permutations. It also should beunderstood that the complexity of the array can have an incompleterepresentation of all potential SPE sequences. For instance, octomersthat have Tm's that are much higher or lower than the average Tm of arandom population may be not be desired to be present. Also, octamersthat have self-complementary 3′ and 5′ ends may exhibit poor bindingability. When more than one species of UPE is being used, this aspectcan be carried out with amplification carried out simultaneously witheach UPE. More preferably, reactions are carried out in parallel with agiven UPE on an array for each set of reactions.

[0238] In another aspect of the present invention, a mixed phaseamplification is carried out where SPE's at fixed locations on an arrayare used for 1^(st) strand synthesis. but the primers used for synthesisof 2^(nd) strands are not attached to the matrix of the array. In thisaspect of the present invention, a pool of primers for 2^(nd) strands insolution can make use of normal nucleic acid kinetics to find 1^(st)strand templates fixed to distinct loci on an array for 2^(nd) strandpriming events.

[0239] FIGS. 22-25 show an example of a series of binding and extensionreactions with only the SPE's fixed to an array. In this example, SPE-P1is a primer fixed to Locus P that is complementary to the (+) strand oftarget P and P2 is a primer that is free in solution and iscomplementary to the (−) strand of target P. SPE-Q1 is a primer fixed toLocus Q that is complementary to the (+) strand of target Q and Q2 is aprimer that is free in solution and is complementary to the (−) strandof target Q.

[0240] It can be seen in FIGS. 22-25 that the specificity of thereaction and anchoring of the amplification to a specific locus can beentirely directed by this 1^(st) strand copying reaction. As such, theidentity of the primers that are free in solution are not important aslong as they are capable of synthesizing nucleic acids that canspecifically bind to the SPE's on the array. Thus although, uniquespecific sequences were used in FIGS. 22-25 for illustration of 2^(nd)strand priming/extension reactions, in this aspect of the inventionwhere a mixed phase amplification is carried out, the primers forsynthesis of 2^(nd) strands could also be a carried out by a mixture ofUPE's or they can even comprise a pool of or random primers. Thisparticular aspect of the present invention also finds use with generalarrays that represent multitudes of variations of sequences. Forinstance, an array that is created by in situ synthesis as described byAffymatrix can be synthesized with some or all of the 65,536permutations of an octamer array and then used in conjunction with UPE'sin solution.

[0241] Another aspect of the present invention discloses novel methods,compositions and kits for the preparation and use of protein and ligandarrays which serve to increase the exposure of the binding substance onthe array and decrease non-specific binding to the matrix itself. In oneembodiment, chimeric compositions are disclosed that are comprised oftwo segments, a nucleic acid portion and a non-nucleic portion. Thenucleic acid portion is used to achieve a practical and more accessiblemethod for attaching the non-nucleic acid portion to a solid support. Inone method of use, the nucleic acid portion is directly bound to thesurface of the array where it serves as a linker between the arraysurface and the non-nucleic acid portions of the chimeric compositions.In addition, due to the phosphate charges of the nucleic acid, eachchimeric composition at a locus should exhibit repulsive forces thatshould minimize interactions between the chimeric compositions.

[0242] Since use is being made of its physical properties rather thanits sequence identity, any particular sequence can be used genericallyfor all the various chimeric compositions. Information on the identityof the non-nucleic acid portion is not derived from the nucleic acidportion but rather form the spatial location on the array where thechimeric composition has been fixed or immobilized. This is in contrastto prior art, which intrinsically required a diversity of specificsequences for the nucleic acid portion and a subsequent “decoding” ofthe nucleic acid portion. In another embodiment of the presentinvention, the nucleic acid portion of the chimeric compositioncomprises discrete sequences that allow binding of the chimericcomposition to the array through hybridization to complementarysequences that are immobilized on the support.

[0243] The nucleic acid portion of a chimeric composition can becomprised of deoxynucleotides, ribonucleotides, modified nucleotides,nucleic acid analogues such as peptide nucleic acids (PNAs), or anycombination thereof. The sequence of the nucleic acid portion is ofcompletely arbitrary nature and may be chosen by the user. In one aspectof the present invention, advantage is taken of the intrinsic propertiesof nucleic acid hybridization for the attachment of the non-nucleic acidportion to the solid surface used for the array. Thus, the presentinvention allows the high specificity, tight binding and favorablekinetics that are characteristic of nucleic acid interactions to beconveyed to a non-nucleic acid portion that does not enjoy theseproperties.

[0244] The non-nucleic acid portion of the chimeric composition of thepresent invention can be comprised of peptides, proteins, ligands or anyother compounds capable of binding or interacting with a correspondingbinding partner. Peptides and proteins can be comprised of amino acidsequences ranging in length from small peptides to large proteins. Thispeptides and proteins can also comprise modified amino acids oranalogues of amino acids. The amino acids or analogues can comprise anydesirable sequence. For instance, the amino acid sequences can bederived from enzymes, antibodies, antigens, epitopes of antigens,receptors and glycoproteins. When peptides or proteins are used as thenon-nucleic acid portion of the chimeric composition, the sequences ofthe nucleic acid portion are of arbitrary nature and have nocorrespondence to the amino acid sequences of the peptides or proteins.Other molecules besides peptides and proteins may also find use in thepresent invention. Examples of other constituents that could be used forthe non-nucleic acid portion can comprise but not be limited to ligandsof MW of 2000 or less, substrates, hormones, drugs and any possibleprotein binding entity.

[0245] As described previously, the particular sequence of the nucleicacid is determined by the user. In one method of use of the presentinvention, each individual species that is used as the non-nucleic acidportion can be covalently joined to a unique nucleic acid sequence.Hybridization of a the nucleic acid portion of the chimeric compositionto a complementary sequence at a particular locus on an array therebydetermines the identity of the particular species of the non-nucleicacid portion that is now bound to that locus. For example, one hundreddifferent chimeric compositions can be synthesized that each comprises aunique peptide and a unique nucleic acid sequence. Hybridization canthen be carried out with an array that has one hundred different loci,where each locus has nucleic acids complementary to one of the uniquenucleic acid sequences. Hybridization thereby results in thelocalization of each unique peptide to one particular locus on thearray, transforming a nucleic acid array into a peptide array. A usefulmethod for selection of sequences that could be used for the nucleicacid portion has been described by Hirschhorn et al., (op.cit.) herebyincorporated by reference. Also, since no relationship is requiredbetween the non-nucleic portion and the nucleic acid portion, adifferent set of one hundred chimeric compositions can be designed thathave different species used for the non-nucleic acid portion but use thesame set of one hundred sequences for the nucleic acid portion. In thisway, a generic nucleic acid array can be used to create differentpeptide arrays by changing the identities of the chimeric compositions.

[0246] Alternatively, non-nucleic acid protein binding substances can beattached to oligonucleotides which all comprise the same sequence. Forexample, chimeric compositions with various non-nucleic portions couldbe synthesized where the nucleic acid portion of each chimericcompositions comprised a common poly T sequence. The matrix can beprepared so that the oligonucleotides at each site consist ofcomplementary Poly A sequences. The chimeric compositions can then beapplied to the matrix using an addressable arraying system that has beendescribed by Heller et al. in U.S. Pat. No. 5,605,662 (hereinincorporated by reference). By these means, each particular chimericcomposition can be applied individually to the matrix using anelectronically controlled system and immobilized through hybridizationto the appropriate site.

[0247] The chimeric compositions at a particular locus of an array donot have to be completely uniform in nature, i.e. an oligonucleotidesequence can be attached to several different species of non-nucleicacid portions. For example, a series of one hundred peptides can beplaced on the array in only four different sites by making Pool 1 withtwenty-five peptides conjugated to oligonucleotide 1, Pool 2 withtwenty-five peptides conjugated to oligonucleotide 2, Pool 3 withtwenty-five peptides conjugated to oligonucleotide 3 and Pool 4 withtwenty-five peptides conjugated to oligonucleotide 4. Attachment of thevarious pools of chimeric compositions to each locus can be carried outby having oligounucleotide 1, 2, 3 and 4 comprising unique sequencescomplementary to different oligonucleotides at each site or as describedabove, an addressable arraying system can be used to localize each poolusing nucleic acid portions with identical sequences. The chimericcompositions comprised of nucleic acid and non-nucleic acid portions canbe synthesized using any method known to those skilled in the art.Methods that may find use with the present invention are described in areview by Tung, C.-H.;(2000 Bioconjugate Chemistry 11, 5, 605-618) andEngelhardt et al., U.S. Pat. No. 5,241,060, issued Aug. 31, 1993 andPergolizzi et al., U.S. patent application Ser. No. 08/479,995, filedJun. 7, 1995, for Analyte Detection Utilizing Polynucleotide Sequences,Composition, Process and Kit, based on priority U.S. patent applicationSer. No. 06/491,929, filed May 5, 1983, all incorporated herein byreference. In one approach, peptides and oligonucleotides aresynthesized separately using standard automated procedures and thencovalently bonded together. For example, a thiol group can be addedeither to the 5′-terminus of the oligonucleotide or internally in thenucleic acid portion of the chimeric composition. Addition of amaleimido group to the N-terminus or in an internal position of thepeptide allows a reaction with the thiol group of the oligonucleotide toform a chimeric composition comprised of a nucleic acid and a peptide(Eritja et al., (1991) Tetrahedron, 47; 4113-4120. Arar et al.; (1993)Tetrahedron Lett 34; 8087-8090, Ede et al., (1994) BioconjugateChemistry 5; 373-378, Stetsenko and Gait, (2000) J. Org. Chem. 65;4900-4908). Alternatively the chimeric composition can be prepared bythe stepwise addition of amino acids and nucleotides on the same solidsupport, (de la Torre et al., (1994) Tetrahedron Lett 35; 2733-2736,.Bergmann and Bannwarth (1995) Tetrahedron Lett. 36; 1839-1842, Robles etal., (1999) Tetrahedron 55; 13,251-13,264, Antopolsky et al., (1999)Helv. Chim Acta 82; 2130-2140). In these publications each of which isincorporated by reference herein, the peptide was synthesized firstfollowed by the addition of bases to synthesize the oligonucleotideportion. In standard peptide synthesis, the N-terminus and the sidechains of the amino acids are protected by Fmoc and tert-butyl groupsrespectively. At each cycle the Fmoc group is removed with 20%piperidine and the side chains are deprotected with 90% trifluoroaceticacid. However when both oligonucleotides and peptides were synthesizedas part of a single composition, different chemistries had to be used.For example, base labile Fmoc and 9-fluorenylmethyl groups were used asthe amino acid side chain protecting groups to avoid exposing the DNA tostrong acids (de la Torre, op cit.; de la Torre et. al., 1999Bioconjugate Chem.10; 1005-1012; Robles et al op cit.), all suchpublications being incorporated by reference herein. Methods for makingchimeric compositions of peptides fused to PNA analogues of nucleicacids have been described by Cook et al. in U.S. Pat. No. 6,204,326,incorporated herein by reference. Furthermore, chimeric compositionscomprised of nucleic acids and peptides can be synthesized directly on asolid surface to create an array using the methods described by Sundberget al in U.S. Pat. No. 5,919,523 incorporated herein by reference.

[0248] The solid support can be any material used for arrays including,but not limited to nylon or cellulose membranes, glass, synthetic,plastic, metal. The materials can be opaque, reflective, transparent ortranslucent. They can be porous or they can be non-porous. Nucleic acidsthat are either part of chimeric compositions or meant to becomplementary to chimeric compositions can be affixed to the solidsupport by any previously known methods used to prepare DNA arrays.

[0249] Binding of analytes to appropriate binding partners can becarried out in either a mixed phase or a liquid phase format. Forinstance, the present invention has disclosed the direct fixation ofbinding substances to the array by the use of rigid arm linkers andchimeric compositions. The binding substance on the array (the solidphase) can be exposed to a solution (the liquid phase) that contains theanalytes of interest. Interactions between the binding substance on thearray and analytes in solution can then later be quantified. Examples ofthe interactions that may find use in the present invention can comprisebut not be limited to peptide-protein, antigen-antibody, ligand-receptoror enzyme-substrates. For example, an array can be prepared with aseries of peptides to determine their ability to bind to a particularantibody. The array is incubated in a solution containing the antibodyfollowed by washing away the unbound material. Detection of the antibodybound to various components on the array can then be carried out by anyof a number of conventional techniques. For instance, in this examplethe antibody that is applied to the array can be labeled with biotin forindirect detection, or a fluorescent compound for direct detection.Alternatively, the antibody analyte is unlabeled and a secondaryantibody can be utilized which either has a fluorescent label for directdetection or indirect label such as biotin. Thus, in this example theantibody-antigen interaction occurs with the antigen bound to the solidmatrix.

[0250] The present invention has also disclosed the use of chimericcompositions that are indirectly bound to the array throughhybridization of the nucleic acid portions of the chimeric compositionsto complementary nucleic acids fixed or immobilized to the array. Thesecan be used in the in the same mixed phase format that has beendescribed above by hybridization of the chimeric compositions to thearray followed by binding of analytes. However, the use of hybridizationto immobilize the chimeric compositions to specific loci on the arrayallows the use of a completely liquid phase format for binding ofanalytes to the chimeric compositions. In this way, the chimericcompositions can be combined with the target molecules in solution underoptimal conditions for interactions between the analyte and thenon-nucleic acid portions of the chimeric compositions. The resultantsolution, containing the chimeric compositions free in solution as wellas the chimeric compositions that are bound into complexes with theanalytes, can then be applied to the matrix and the various chimericcompositions will be localized to various locations on the array throughhybridization to the nucleic acid portion to complementary sequences onthe array. An illustration of this process is given in FIG. 28.

[0251] The hybridization can be carried out under mild conditions, whichwill not interfere with the ligand-receptor or protein-protein complex.Protein-protein interactions are generally characterized by low Km's, inthe order of magnitude of 10⁻⁵ to 10⁻⁹. In this aspect of the presentinvention, the protein interactions can occur in solution rather than ona solid surfaces which will allow superior kinetics of binding and willalso allow a wider variety of conditions for protein binding than can beobtained in the mixed format. Also, by chimeric compositions andanalytes together in solution, direct interaction or interference withthe matrix is avoided, thereby decreasing the background. Therefore, touse the example cited before, the solution containing the antibodytarget is combined with a solution containing the chimeric composition.Thus, by using the methods of the present invention, the proteins willremain in solution throughout the process preventing any problemsassociated with dehydrating the protein bound to the solid matrix.

[0252] The method of the present invention can be used to study manysystems that involve interactions between compound. These can includebut not be limited to antigen-antibody relationships, protein-proteininteractions, enzyme-substrate receptor-ligand interactions,ligand-receptor, hormone-receptor, carbohydrate-lectins, drug screening,and patterns of expression of proteins in a cell or tissue. Anothermethod of use of the present invention is that instead of using uniquenucleic acid portions for each individual non-nucleic acid portion, onespecific binding substance can be combined with various nucleic acidsources to form a group of chimeric compositions with a commonnon-nucleic acid portion and a unique nucleic acid portion. Eachparticular chimeric composition can be combined with an analyte from adifferent source and applied to the array by hybridizing the nucleicacid portions to their complementary sequences on the array. Theproteins bound to the array can then be detected following standardprocedures. By these means, the amount of targets from each source thatcan interact with the binding substance in the chimeric compositions canbe simultaneously determined.

[0253] For instance, a set of twenty different compositions can besynthesized where each member of the set will have a different nucleicacid portion but the same peptide. Another set can be made with adifferent peptide that is linked to twenty other nucleic acid portions.More sets can be made on the same basis. Protein extracts can then bemade from twenty different tissues and each extract can be combined witha different member of the set of chimeric compositions. Thus, thenucleic acid portion serves as a marker for not only the peptide butalso for the particular tissue that was used as the source. Forinstance, a group of sets can be made with peptides that have affinitiesfor different receptors. After incubation of the mixtures with thechimeric compounds, the mixtures are applied to the array and detected.In this way, each particular receptor that is being studied can bequantified and compared simultaneously between various tissues.Alternatively, the same nucleic acid sequence can be used in common foreach source by using the addressable system described previously, andcarrying out hybridization to each locus after addition of eachindividual reaction mixture.

[0254] The same method can be applied to tissues or cell cultures thatare from the same source but are treated differently. For example, in adrug discovery program, nine different drugs can be added to individualcell cultures to determine the effect on specific proteins. Chimericcompositions are designed and synthesized with peptides that are knownto react with each of proteins that is to be monitored. As in theprevious example, a specific nucleic acid sequence will serve as amarker for each peptide and each particular treatment. The proteins areextracted from each of the ten cell cultures (nine drug treated plus anuntreated control) and incubated with the chimeric compositions. Themixtures are applied to the array and the amount of analyte bound to thecorresponding peptides at each locus of the array is measured for thevarious drug conditions. If desired, the present invention can also beused for the isolation of analytes. This can be carried out by eitherdisrupting the interaction between the analyte and the non-nucleic acidportion of the chimeric compositions or by denaturing the nucleic acidportion from the complementary sequence fixed or immobilized to thearray. It is also contemplated that removal of chimeric compositionsfrom the array may also allow the reuse of the array in otherexperiments.

[0255] In further detail, this invention provides novel chimericcompositions and processes using such chimeric compositions. One suchcomposition of matter comprises an array of solid surfaces comprising aplurality of discrete areas, wherein at least two of such discrete areascomprise: a chimeric composition comprising a nucleic acid portion; anda non-nucleic acid portion, wherein the nucleic acid portion of a firstdiscrete area has the same sequence as the nucleic acid portion of asecond discrete area, and wherein the non-nucleic acid portion has abinding affinity for analytes of interest.

[0256] Another composition of matter comprises an array of solidsurfaces comprising a plurality of discrete areas; wherein at least twoof the discrete areas comprise a chimeric composition hybridized tocomplementary sequences of nucleic acids fixed or immobilized to thediscrete areas, wherein the chimeric composition comprises a nucleicacid portion, and a non-nucleic acid portion, the nucleic acid portioncomprising at least one sequence, wherein the non-nucleic acid portionhas a binding affinity for analytes of interest, and wherein when thenon-nucleic acid portion is a peptide or protein, the nucleic acidportion does not comprises sequences which are either identical orcomplementary to sequences that code for such peptide or protein.

[0257] Mention should be made of a process for detecting or quantifyinganalytes of interest, the process comprising the steps of 1) providinga) an array of solid surfaces comprising a plurality of discrete areas,wherein at least two of such discrete areas comprise a chimericcomposition comprising a nucleic acid portion, and a non-nucleic acidportion; wherein the nucleic acid portion of a first discrete area hasthe same sequence as the nucleic acid portion of a second discrete area;and wherein the non-nucleic acid portion has a binding affinity foranalytes of interest; b) a sample containing or suspected of containingone or more of the analytes of interest; and c) signal generating means;2) contacting the array a) with the sample b) under conditionspermissive of binding the analytes to the non-nucleic acid portion; 3)contacting the bound analytes with the signal generating means; and 4)detecting or quantifying the presence of the analytes.

[0258] Another process for detecting or quantifying analytes of interestcomprises the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of suchdiscrete areas comprise a chimeric composition comprising a nucleic acidportion; and a non-nucleic acid portion; wherein the nucleic acidportion of a first discrete area has the same sequence as the nucleicacid portion of a second discrete area; and wherein the non-nucleic acidportion has a binding affinity for analytes of interest; b) a samplecontaining or suspected of containing one or more of the analytes ofinterest; and c) signal generating means; 2) labeling the analytes ofinterest with the signal generating means; 3) contacting the array a)with the labeled analytes under conditions permissive of binding thelabeled analytes to the non-nucleic acid portion; and 4) detecting orquantifying the presence of the analytes.

[0259] Another process for detecting or quantifying analytes of interestcomprises the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of suchdiscrete areas comprise nucleic acids fixed or immobilized to suchdiscrete areas, b) chimeric compositions comprising: i) a nucleic acidportion; and ii) a non-nucleic acid portion; the nucleic acid portioncomprising at least one sequence, wherein the non-nucleic acid portionhas a binding affinity for analytes of interest, and wherein when thenon-nucleic acid portion is a peptide or protein, the nucleic acidportion does not comprise sequences which are either identical orcomplementary to sequences that code for the peptide or protein; c) asample containing or suspected of containing the analytes of interest;and d) signal generating means; 2) contacting the array with thechimeric compositions to hybridize the nucleic acid portions of thechimeric compositions to complementary nucleic acids fixed orimmobilized to the array; 3) contacting the array a) with the sample b)under conditions permissive of binding the analytes to the non-nucleicacid portion; 4) contacting the bound analytes with the signalgenerating means; and 5) detecting or quantifying the presence of theanalytes.

[0260] Another process for detecting or quantifying analytes of interestcomprises the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of thediscrete areas comprise nucleic acids fixed or immobilized to thediscrete areas, b) chimeric compositions comprising i) a nucleic acidportion; and ii) a non-nucleic acid portion, the nucleic acid portioncomprising at least one sequence, wherein the non-nucleic acid portionhas a binding affinity for analytes of interest, and wherein when thenon-nucleic acid portion is a peptide or protein, the nucleic acidportion does not comprise sequences which are either identical orcomplementary to sequences that code for the peptide or protein; c) asample containing or suspected of containing the analytes of interest;and d) signal generating means; 2) contacting the chimeric compositionswith the sample b) under conditions permissive of binding the analytesto the non-nucleic acid portion; 3) contacting the array with thechimeric compositions to hybridize the nucleic acid portions of thechimeric compositions to complementary nucleic acids fixed orimmobilized to the array; 4) contacting the bound analytes with thesignal generating means; and 5) detecting or quantifying the presence ofthe analytes.

[0261] Another useful process comprises the steps of 1) providing a) anarray of solid surfaces comprising a plurality of discrete areas;wherein at least two of the discrete areas comprise nucleic acids fixedor immobilized to the discrete areas, b) chimeric compositionscomprising i) a nucleic acid portion; and ii) a non-nucleic acidportion; the nucleic acid portion comprising at least one sequence,wherein the non-nucleic acid portion has a binding affinity for analytesof interest, and wherein when the non-nucleic acid portion is a peptideor protein, the nucleic acid portion does not comprise sequences whichare either identical or complementary to sequences that code for thepeptide or protein; c) a sample containing or suspected of containingthe analytes of interest; and d) signal generating means; 2) contactingthe array with the chimeric compositions to hybridize the nucleic acidportions of the chimeric compositions to complementary nucleic acidsfixed or immobilized to the array; 3) labeling the analytes of interestwith the signal generating means; 4) contacting the array with thelabeled analytes to bind the analytes to the non-nucleic acid portion;and 5) detecting or quantifying the presence of the analytes.

[0262] Another process for detecting or quantifying analytes of interestcomprises the steps of 1) providing a) an array of solid surfacescomprising a plurality of discrete areas; wherein at least two of thediscrete areas comprise nucleic acids fixed or immobilized to thediscrete areas, b) chimeric compositions comprising: i) a nucleic acidportion; and ii) a non-nucleic acid portion; the nucleic acid portioncomprising at least one sequence, wherein the non-nucleic acid portionhas a binding affinity for analytes of interest, and wherein when thenon-nucleic acid portion is a peptide or protein, such nucleic acidportion does not comprise sequences which are either identical orcomplementary to sequences that code for the peptide or protein; c) asample containing or suspected of containing the analytes of interest;and d) signal generating means; 2) contacting the array with thechimeric compositions to hybridize the nucleic acid portions of thechimeric compositions to complementary nucleic acids fixed orimmobilized to the array; 3) labeling the analytes of interest with thesignal generating means; 4) contacting the array with the labeledanalytes to bind the analytes to the non-nucleic acid portion; and 5)detecting or quantifying the presence of the analytes.

[0263] The elements recited in the last several chimeric compositionsand processes using such chimeric compositions are described elsewherein this disclosure.

[0264] The examples which follow are set forth to illustrate variousaspects of the present invention but are not intended in any way tolimit its scope as more particularly set forth and defined in the claimsthat follow thereafter.

DESCRIPTION OF THE PREFERRED EMBODIMENTS EXAMPLE 1 Amplification of aLibrary of RNA Targets with 2^(nd) Strand Synthesis Carried Out byRandom Primers with T7 Promoter Sequences

[0265] 1) First Strand Synthesis

[0266] Two mixtures of 250 ng of rabbit globulin mRNA (LifeTechnologies, Rockville, Md.) and 200 ng of Oligo (dT)₂₄ (In house orpurchased?) in 5 ul were heated at 70° C. for 10 minutes followed by a 2minute incubation on ice. This material was then used in 10 ul reactionscontaining 50 mM Tris-HCl (pH 8.3), 75 mM KCl, 3 mM MgCl₂, 10 mM DTT,600 uM dNTPs and 120 units of Superscript II RNase H⁻ ReverseTranscriptase (Life Technologies, Rockville, Md.) with incubation at 42°C. for 60 minutes.

[0267] 2) Second Strand Synthesis

[0268] KOH was added to the reactions for a final concentratiion of 200mM. Incubation was carried out at 37° C. for 30 minutes followed byneutralization with an equimolar amount of glacial acetic acid. Primerswith the following sequence were used for 2^(nd) strand synthesis:

[0269] 5′-GGCCAGTGAATTGTAATACGACTCACTATAGGGAGGCGGN₁₋₉-3′

[0270] Primers with the sequence above (TPR primers) consist of a T7promoter sequence at their 5′ ends and 9 nucleotides with randomsequences at their 3′ ends. 400 pmoles of TPR primers and otherappropriate reagents were added for a final reaction mix of 30 ulcontaining 86.6 mM Tris-HCl (pH 7.6), 32 mM KCl, 200 mM KOAc (??), 15.6mM MgCl₂, 3.3 mM DTT, 10 mM Dithioerythritol (DTE), 10 mM (NH₄)₂SO₄,0.15 mM −NAD, 200 ug/ml nuclease-free BSA (Bayer, Kankakee, Ill.),Annealing was carried out by heating the mixture to 65° C. and slowcooling to room temperature followed by incubation on ice for 5 minutes.Extension of the primers was carried out by addition of 1.2 ul of 10 mMdNTPs, 4 units of E. coli DNA ligase (New England Biolabs, Beverly,Mass.) and either 12 units of DNA polymerase I (New England Biolabs,Beverly, Mass.) or 6 units of the Exo (−) version of the Klenow fragmentof DNA Polymerase I (New England Biolabs, Beverly, Mass.). Incubationwas carried out at 15° C. for 5 minutes followed by 37° C. for 120minutes. The reactions were puriifed by extraction withPhenol/Chloroform with Phase-Lock Gels(Eppendorf, Westbury, N.Y.) andEthanol precipitated.

[0271] 3) Transcription

[0272] Transcription was carried out by using the BioArray High YieldTranscription Kit (T7) (ENZO Diagnostics, Farmingdale, N.Y.) followingthe manufacturers instructions with a final volume of 40 ul. Thereaction mixes also contained 10 uCi of ³H-ATP with a specific activityof 45 Ci/mMol (Amersham Pharmacia, Piscataway, N.J.). Incorporation wasmeasured by addition of 5 ul of the transcription reaction to 1 ml of10% TCA, 50 ug/ml Poly A, 5 mM EDTA followed by incubation on ice for 30minutes. Precipitates were collected on 25 mm glass fiber filters(Whatman, Lifton, N.J.) followed by three washes with 5% TCA and threewashes with ethanol

[0273] 4) Results and Conclusions Sample 1 with DNA polymerase I  4,243cpm Sample 2 with Exo (−) Klenow 19,662 cpm

[0274] This example demonstrated that RNA transcripts were obtained froma library of nucleic acids by the steps described above and that underthe conditions used, the Exo (−) version of Klenow resulted in moreproduct compared to the use of DNA polymerase 1.

EXAMPLE 2 Amplification of a Library of RNA Targets with 1^(st) StrandSynthesis Using Oligo-T Magnetic Beads and 2^(nd) Strand SynthesisCarried Out by Random Primers with T7 Promoter Sequences

[0275] 1) Preparation of Beads

[0276] 50 ul of Dynal Oligo (dT)₂₅ magnetic beads (Dynal Inc., LakeSuccess, N.Y.) were washed two times with 100 ul of Binding Buffer (20mM Tris-HCl (pH 7.5), 1.0 M LiCl, 2 mM EDTA) and then resuspended in 50ul of Binding Buffer.

[0277] 2) Binding of RNA to Beads

[0278] RNA targets were prepared by diluting I ug of mouse poly A RNA(Sigma Chemical Co, St. Louis, Mo.) or 1 ug of wheat germ tRNA (SigmaChemical Co, St. Louis, Mo.) into RNase-free H₂O (Ambion, Austin, Tex.)for a final volume of 50 ul, and heating the RNA solution at 65° C. for5 minutes. The RNA solution was combined with the beads prepared in Step1 and mixed for 15 minutes at room temperature with a Dynal Sample Mixer(Dynal Inc., Lake Success, N.Y.). Unbound material was removed bymagnetic separation with a Dynal Magnetic Particle Concentrator (Dynal,Inc. Lake Success, N.Y.) followed by two washes with 200 ul of WashBuffer B (10 mM Tris-HCl (pH 7.5), 150 mM LiCl, 1 mM EDTA) and threewashes with 250 ul of First Strand Buffer (50 mM Tris-HCl (pH 8.3), 75mM KCl, 3 mM MgCl₂)

[0279] 3) First Stand Synthesis

[0280] The beads from Step 2 were resuspended in 50 mM Tris-HCl (pH8.3), 75 mM KCl, 3 mM MgCl₂, 10 mM DTT, 500 uM dNTPs and 400 units ofSuper Script II RNase H⁻ Reverse Transcriptase (Life Technologies,Rockville, Md.) and incubated for 90 minutes at 42° C.

[0281] 4) Second Strand Synthesis

[0282] RNA templates were removed by heating the First Strand Synthesisreaction mixture of step 3 at 90° C. for 5 minutes followed by removalof the supernatant after magnetic separation. The beads were washed twotimes with 100 ul of Buffer C (70 mM Tris-HCl (ph 6.9) 90 mM KCl, 14.6mM MgCl₂, 10 mM DTE, 10 mM (NH₄)₂SO4 and 200 ug/ml nuclease-free BSA)and resuspended in 50 ul of Random Priming Mix A (86.7 mM Tris-HCl (pH7.6), 113.3 mM KCl, 17 mM MgCl₂, 11.3 mM DTT, 11.3 mM (NH₄)₂SO₄, 227ug/mi nuclease-free BSA) containing 360 pmoles of TPR primers. Primerswere allowed to anneal on ice for 15 minutes. Unbound primers wereremoved by magnetic separation. The beads were resuspended in 50 ul ofRandom Priming Mix A (without the TPR primers) with 10 units of theKlenow fragment of DNA Polymerase I (New England Biolabs, Beverly,Mass.) and 400 mM dNTP's. Incubation was carried out for 5 minutes at 4°C., 30 minutes at 15° C., and 30 minutes at 37° C. For some samples, anadditional 25 ul of Oligo T magnetic beads prepared as described in Step1 were washed with Buffer C and added to the reaction mix. Also, forsome samples, 3 units of T4 DNA Polymerase (New England Biolabs,Beverly, Mass.) and 2 ul of a 10 mM stock of dNTPs were added to thereaction mixtures. Samples with these further steps were incubated for30 minutes at 37° C. At the conclusion of the varied reactions, thebeads were magnetically separated from the reagents.and the beads wereused to carry out transcription assays.

[0283] 5) Transcription Synthesis

[0284] Transcription reactions were carried out by resuspending thebeads in reagents from the BioArray High Yield Transcription Kit (T7)(ENZO Diagnostics, Farmingdale, N.Y.) using the manufacturer'sinstructions with a final volume of 40 ul. The reaction mixtures alsocontained 10 uCi of ³H-ATP with a specific activity of 45 Ci/mMol(Amersham Pharmacia, Piscataway, N.J.). Extent of transcription wasmeasured by using TCA precipitation as described previously.

[0285] 6) Results Extra T4 DNA Sample Target Beads polymerase cpmIncorporated 1 Poly A (−) (−) 8,535 2 Poly A (−) (+) 15,483 3 Poly A (+)(−) 16,048 4 Poly A (+) (+) 18,875 5 tRNA (+) (+) 2,548

[0286] 7) Conclusions

[0287] This example demonstrated that transcripts were obtained from alibrary of nucleic acids by the steps described above. Addition of extrabeads can increase the amount of synthesis. The reaction can be carriedout without a T4 DNA polymerization step but the amount of synthesis canbe increased by the addition of such a reagent.

EXAMPLE 3 Dependency on Reverse Transcriptase for Amplification of aLibrary of RNA Targets with Oligo-T Magnetic Beads and Random Primerswith T7 Promoter Sequences

[0288] 1) Preparation of Beads

[0289] This step was carried out as described in Step 1 of Example 2,except the amount of beads was increased to 100 ul for each reaction

[0290] 2) Binding of RNA to Beads

[0291] RNA targets were prepared by diluting I ug of mouse poly A mRNA(Sigma Chemical Co, St. Louis, Mo.) into nuclease-free H₂O (Ambion Inc.,Auistin Tex.) for a final volume of 50 ul , and heating the RNA solutionat 65° C. for 15 minutes. The RNA solution was combined with the beadsprepared in Step 1 and mixed for 15 minutes at Room Temperature with aDynal Sample Mixer (Dynal Inc., Lake Success, N.Y.). Unbound materialwas removed by magnetic separation followed by two washes with 200 ul ofWash Buffer B and two washes with 100 ul of First Strand Buffer.

[0292] 3) First Strand Synthesis

[0293] This step was carried out as described in step 3 of Example 2except that a pair of duplicate samples had the Reverse Transcriptaseomitted

[0294] 4) Second Strand Synthesis

[0295] RNA templates were removed by heating the First Strand Synthesisreaction mixture of step 3 at 90° C. for 4 minutes followed by removalof the supernatant after magnetic separation. The beads were washed twotimes with 100 ul of Wash Buffer B and resuspended in 50 ul of RandomPriming Mix A containing 360 pmoles of TPR primers. Primers were allowedto anneal on ice for 15 minutes. Unbound primers were removed bymagnetic separation and the beads were washed twice with 100 ul of coldBuffer D (20 mM Tris-HCl (pH 6.9), 90 mM KCl, 4.6 mM MgCl₂, 10 mM(NH₄)₂SO₄. The beads were then suspended in 40 ul of Buffer C that alsocontained 1 mM dNTPs and 10 units of the Klenow fragment of DNAPolymerase I (New England Biolabs, Beverly, Mass.). Incubation wascarried out for 5 minutes at 4° C., 30 minutes at 15° C., and 30 minutesat 37° C. The reaction was carried out further by the addition of 2 ul(6 units) of T4 DNA Polymerase (New England Biolabs, Beverly, Mass.) and2 ul of a 10 mM stock of dNTPs, followed by incubation for 30 minutes at37° C.

[0296] 5) Transcription Synthesis

[0297] The beads were washed two times with 100 ul of Wash Buffer B andonce with 100 ul of 10 mM Tris-HCl (pH 7.5). The beads were resuspendedin 10 ul of 10 mM Tris-HCl (pH 7.5) and mixed with reagents from aBioArray High Yield Transcription Kit (T7) (ENZO Diagnostics,Farmingdale, N.Y.) using the manufacturer's instructions. The volume ofthe reaction was 30 ul and the incubation was carried out for 2 hours at37° C.

[0298] 6) Results and Conclusions

[0299] Analysis of the reaction was carried out by gel electrophoresisof 10 ul of the transcription reaction using 1% Agarose in 0.5×TBEbuffer. The results of this experiment are in FIG. 27 for duplicatesamples and demonstrate that transcripts were obtained from a library ofnucleic acids by the steps described above and this synthesis wasdependent upon the presence of Reverse Transcriptase activity.

EXAMPLE 4 Multiple rounds of Synthesis of 2^(nd) Strands by RandomPrimers with T7 Promoters

[0300] Steps 1, 2 and 3 for Preparation of beads, binding of mRNA and1^(st) strand synthesis were carried out as described in steps 1 through3 of Example 3.

[0301] 4) Second Strand Synthesis

[0302] After 1^(st) strand synthesis, the liquid phase was removed bymagnetic separation and the beads resuspended in 100 ul of DetergentWash No.1 (10 mM Tris-HCl (pH 7.5), 1 % SDS) and heated at 90° C. for 5minutes.. The supernatant was removed by magnetic separation and thebeads were washed with 100 ul of Detergent Wash No.2 (40 mM Tris-HCl (pH8.0), 200 mM KCl, 0.2 mM EDTA, 0.01% Tween 20, 0.01% Nonidet P4O). Thebeads were washed two times with 100 ul of Wash Buffer B and resuspendedin 50 ul of Random Priming Mix A containing 360 pmoles of TPR primers.Primers were allowed to anneal on ice for 15 minutes. Unbound primerswere removed by magnetic separation and the beads were washed twice with100 ul of cold Buffer D (20 mM Tris-HCl (pH 6.9), 90 mM KCl, 4.6 mMMgCl₂, 10 mM DTT, 10 mM (NH₄)₂SO4). The beads were then suspended in 40ul of Buffer C that also contained 1 mM dNTPs and 10 units of the Klenowfragment of DNA Polymerase I (New England Biolabs, Beverly, Mass.).Incubation was carried out for 5 minutes at 4° C., 30 minutes at 15° C.,and 30 minutes at 37° C. The reaction was carried out further by theaddition of 2 ul (6 units) of T4 DNA Polymerase (New England Biolabs,Beverly, Mass.) and 2 ul of a 10 mM stock of dNTPs, followed byincubation for 30 minutes at 37° C. The beads were then washed two timeswith 100 ul of Wash Buffer B, resuspended in 50 ul of 10 mM Tris-HCl (pH7.5) and heated at 90° C. for 5 minutes. The supernatant was removedafter magnetic separation and store as supernatant No.1. The beads werethen washed once with 100 ul of Detergent Wash No.2, two times with 100ul of Wash Buffer B and resuspended in 50 ul of Random Priming Mix Acontaining 360 pmoles of TPR primers. Primer annealing and extension wascarried out as described above. The beads were then washed two timeswith 100 ul of Wash Buffer B, resuspended in 50 ul of 10 mM Tris-HCl (pH7.5) and heated at 90° C. for 5 minutes. The supernatant was removedafter magnetic separation and store as supernatant No.2. The series ofwashes, annealing and extension steps were carried out again using thesteps described above. The beads were then washed two times with 100 ulof Wash Buffer B, resuspended in 50 ul of 10 mM Tris-HCl (pH 7.5) andheated at for 5 minutes. The supernatant was removed after magneticseparation and stored as supernatant No.3.

[0303] 5) Synthesis of Complements to the 2^(nd) Strands

[0304] A pool was created by combining supernatant No.1, supernatantNo.2 and supernatant No.3. This pool comprises a library of 2^(nd)strands free in solution with T7 promoters at their 5 ′ ends and poly Asegments at their 3′ ends. Fresh magnetic beads with poly T tails wereprepared and annealed to the pool of 2^(nd) strands by the sameprocesses described in Steps 1 and 2 of Example 2. Extension was thencarried out by resuspension of beads in 50 ul of Buffer C that alsocontained 1 mM dNTPs and 10 units of the Klenow fragment of DNAPolymerase I (New England Biolabs, Beverly, Mass.). Incubation wascarried out at 37° C. for 90 minutes. Transcription was then carried outas described in step 5 of Example 3 except the reaction volume wasreduced to 20 ul.

[0305] 6) Results and Conclusions

[0306] The results of this experiment are in FIG. 28 and demonstratedthat transcripts were obtained from a library of polyA mRNA by the stepsdescribed above. This example demonstrated that a library of 2^(nd)strands was obtained after multiple rounds of 2^(nd) strand synthesis,isolated free in solution and then used to create functionally activeproduction centers

EXAMPLE 5 Additional RNA Synthesis from Transcription Constructs

[0307] The library of transcription constructs described in Example 4were used for a second round of transcription. After removal oftranscription products for analysis in Example 4, the beads wereresuspended in 100 ul of 10 mM Tris-HCl (pH 7.5) and left overnight at4° C. The next day, the beads were washed with 100 ul of Detergent WashNo.2, resuspended in 100 ul of Detergent Wash No.1 and heated at 42° C.for 5 minutes followed by two washes with 100 ul of Detergent BufferNo.2, two washes with 100 ul of Wash Buffer B and two washes with 100 ulof 10 mM Tris-HCl (pH 7.5). A transcription reaction was set up asdescribed previously with a 20 ul volume.

[0308] Results and Conclusions

[0309] Results of the transcription reaction are shown in FIG. 29 andshow that the nucleic acids synthesized in Example 4 were stable andcould be used for additional transcription synthesis.

EXAMPLE 6 Terminal Transferase Addition of Poly G Tail to 1^(st) Strandsfor Binding of Primers with T7 Promoter

[0310] 1) Preparation of Beads

[0311] 150 ul of Dynal Oligo (dT)₂₅ magnetic beads (Dynal Inc., LakeSuccess, N.Y.) were washed two times with 150 ul of Binding Buffer andresuspended in 75 ul of Binding Buffer.

[0312] 2) Binding of RNA to Beads

[0313] RNA targets were prepared by diluting 3 ul of 0.5 ug/ul mousepoly A RNA (Sigma Chemical Co, St. Louis, Mo.) with 32 ul of RNase-freeH₂O (Ambion, Austin, Tex.) and 40 ul of Binding Buffer, and heating theRNA solution at 65° C. for 5 minutes. The RNA solution was combined withthe beads prepared in Step 1 and mixed for 30 minutes at roomtemperature.

[0314] 3) First Strand Synthesis

[0315] Unbound material was removed by magnetic separation followed bytwo washes with 200 ul of Wash Buffer B and one wash with 100 ul ofFirst Strand Buffer. The beads were resuspended in a 50 ul mixture of 50mM Tris-HCl (pH 7.5), 75 mM KCl, 3 mM MgCl₂, 10 mM DTT, 500 uM dNTPs and400 units of Super Script II RNase H⁻ Reverse Transcriptase (LifeTechnologies, Rockville, Md.) and incubated for 90 minutes at 42° C. Atthe end of the 1^(st) strand synthesis reaction, the liquid phase wasremoved by magnetic separation and the beads resuspended in 100 ul ofDetergent Wash No.1 and heated at 90° C. for 5 minutes. The supernatantwas removed by magnetic separation and the beads were washed with 100 ulof Detergent Wash No.2, two times with 100 ul of Wash Buffer B andresuspended in 300 ul of 10 mM Tris-HCl (pH 7.5).

[0316] 4) Second Strand Synthesis

[0317] Two methods were used for carrying out second strand synthesis.The first method was as described for the previous examples, I.e the useof TPR primers that have a T7 promoter on their 5′ ends and randomsequences at their 3′ ends. The second method was the use of T7-C9primers that have a T7 promoter at their 5′ ends and a poly C segment attheir 3′ ends. The sequence of the T7-C9 primers is as follows:

[0318] 5′ GGCCAGTGAATTGTAATACGACTCACTATAGGGATCCCCCCCCC-3′

[0319] The product of Step 3 was divided into two portions. The firstportion (Sample No.1) consisted of 100 ul and was set aside to be usedfor random priming. The second portion (the remaining 200 ul) wasprocessed further by magnetically separating the buffer from the beadsand resuspending the beads in 100 ul and adding 100 ul of Poly A Mix(1.6 ug/ul Poly A, 10 mM Tris-HCL (pH 7.5), 0.5 M LiCl, 1 mM EDTA). ThePoly A was obtained from (Amersham Pharmacia, Piscataway, N.J.) and hadan average length of 350 nucleotides. The beads and Poly A were mixedtogether for 30 minutes at room temperature with a Dynal Sample Mixer(Dynal Inc., Lake Success, N.Y.). The beads were washed two times withWash Buffer B and resuspended in 200 ul of 10 m Tris-HCl (pH 7.5 ). Thiswas divided into two 100 ul portions, Sample No.2 and Sample No.3.Sample No.3 was processed further by magnetically separating the bufferfrom the beads and resuspending the beads in an 80 ul reaction mixtureusing reagents and directions from the 3′ Oligonucleotide Tailing System(ENZO Biochem, Farmingdale, N.Y. 11561) with 0.5 mM dGTP present. SampleNo.3 was incubated for one hour at 37° C. followed by removal of thereagents by magnetic separation. The beads were then resuspended in 100ul of Detergent Buffer No.1 and heated at 90° C. for 3 minutes. Thebeads were then washed once with 100 ul of Detergent Wash No.2 and twicewith 100 ul of Wash Buffer B. Sample No. 3 was resuspended in 100 ul of10 mM Tris-HCl (pH 7.5). All three samples (Sample No.1, Sample No.2 andSample No.3) were washed once with 100 ul Wash Buffer E (100 mM Tris-HClpH7.4) 20 mM KCl, 10 mM MgCl₂, 300 mM (NH4)2S04) and then resuspended in50 ul of Buffer E. Primers for 2^(nd) strand synthesis were added toeach sample: 4 ul of 100 pMole/ul of TPR primers to Sample No.1 and 4 ulof 10 pMole/ul of T7-C9 primers to Samples No.2 and No.3. Samples werethen incubated on ice for 15 minutes followed by one wash with 100 ul ofice cold Buffer E and one wash with ice cold Buffer D. Each sample wasresuspended in 40 ul of Buffer D that also contained 1 mM dNTPs and 200units of the Klenow fragment of DNA Polymerase I (New England Biolabs,Beverly, Mass.). Incubations were carried out for 30 minutes at 15° C.followed by 30 minutes at 37° C.

[0320] All three samples were further processed by the addition of 2 ul(3 units) of T4 DNA polymerase (Source, Location) and 2 ul of 10 mMdNTPs followed by incubation at 37° C. for 30 more minutes. Samples werewashed twice with 100 ul of 10 mM Tris-HCl (pH 7.5). A Transcriptionreaction was set up as described previously with a 20 ul volume.

[0321] 5) Results and Conclusions

[0322] Analysis of the reaction was carried out by gel electrophoresiswith 2 ul and 10 ul samples of the transcription reaction using 1 %Agarose in 0.5×TBE buffer. The results of this experiment are in FIG. 30and demonstrated that non-inherent UDTs were added to the ends of alibrary of 1^(st) strand copies by the methods described above. Thenon-inherrent UDTs served as primer binding sites for primers with polyC at their 3′ ends for synthesis of a library of 2^(nd) strands. Thedifference in the amount of RNA transcription between Samples No.2 andNo. 3 serves as a further indication that comparatively little primingtook place at internal sites under the conditions used.

EXAMPLE 7 Terminal Transferase Addition of Poly G Tail to 1^(st) Strandsfor Binding of Primers with T7 Promoter (Incorporation Assay)

[0323] The transcription products of Example 6 were analyzed by gelelectrophoresis as shown in FIG. 30. To obtain numerical evaluation ofthe method described in that example, the libraries attached to thebeads in Samples No.1, No.2 and No.3 were used in another transcriptionreaction using ³H-incorporation. Transcription was carried out asdescribed in Example 3.

[0324] The results were as follows:

[0325] Random priming

[0326] Sample No.1) 6,660 cpm

[0327] T7-C9 primers without TdT addition step

[0328] Sample No.2 1,144 cpm

[0329] T7-C9 primers with TdT addition step

[0330] Sample No.3 21,248 cpm

[0331] This second assay agrees with the conclusions of Example 6; i.e.the T7-C9 primers can be used in the present method and more primingtook place with the terminally added poly G sequences compared tointernal sequences.

EXAMPLE 8 Incorporation of Promoters After 2^(nd) Strand Synthesis

[0332] 1) Preparation of Beads

[0333] Preparation of beads for each sample was carried out as describedin step 1 of Example 3

[0334] 2) Binding of RNA to Beads

[0335] In each sample, 1 ug of poly mRNA was bound to beads as describedin step 2 of Example 3 with the addition of having 120 units of PrimeRNase Inhibitor (Eppendorf, Westbury, N.Y.) present.

[0336] 3) First Strand Synthesis

[0337] First strand synthesis was carried out as described in step 3 ofExample 3 except the reaction was also supplemented with 120 units ofPrime RNase Inhibitor

[0338] 4) Second Strand Synthesis

[0339] Poly dG addition was carried out as described for sample No. 3 inExample 6. Second strand synthesis was performed as described in Example6 except that 80 pMoles of primers were used in 100 ul reactions. ForSamples No. 1 and No.2, the 2nd strand primers were the T7-C9 primerspreviously described. For Samples No.3 and No.4, the 2nd strand primerswere C9 primers with the sequence: 5′-CCCCCCCCC-3′. At the end of thereaction, all samples were washed twice with 100 ul 10 mM Tris-HCl (pH7.5).

[0340] 5) Third Strand Synthesis

[0341] Samples No.2, No.3 and No.4 were processed further byresuspension of the beads in 26 ul of 10 mM Tris-HCl (pH 7.5) andheating at 90° C. for 3 minutes. The second strands released by thisprocess were isolated apart from the beads by magnetic separation andmixed with 40 pMoles of 3^(rd) strand primers for a final volume of 30ul. For Sample No.3, the 3^(rd) strand primers were T7-T₂₅ primers withthe sequence

[0342] 5′ GGCCAGTGAATTGTAATACGACTCACTATAGGGATC(T)₂₅-3′

[0343] For Samples No.2 and No.4, the 3^(rd) strand primers were T3-T₂₅primers with the sequence:

[0344] 5° CTCAACGCCACCTAATTACCCTCACTAAAGGGAGAT(T)₂₅-3′

[0345] After mixing, Samples No.2, No.3 and No.4 were kept on ice for 15minutes. Extension reactions were then set up in 1×M-MuLV Buffer (NewEngland Biolabs, Beverly Mass.) with 10 units of M-MuILV ReverseTranscriptase (New England Biolabs, Beverly Mass.) and 1 mM of each dNTPin a final volume of 40 ul. Incubation was carried out for one hour at37° C. 6 units of T4 DNA Polymerase (New England Biolabs, Beverly,Mass.) were added to Samples No.1, No.2, No.3 and No.4 and incubationcarried out for a further 15 minutes at 37° C. Reactions were stopped bythe addition of EDTA (pH 8.0) to a final concentration of 10 mM. The DNAfrom Samples 2, No.3 and No.4 was then purified by adjusting the volumesto 150 ul by adding appropriate amounts of 10 mM Tris-HCl. Reactionswere mixed with an equal volume of Phenol:chloroform:isoamyl alcohol(25:24:1) and transferred to 2 ml Phase Lock Gel Heavy tubes(Eppendorf,Westbury, N.Y.). Tubes were vorteed for 1-2 minutes and centrifuged for10 minutes at 16,000 rpm in a microfuge. The aquaeous phase was thentransferred to another tube and DNA precipitated with Ethanol andAmmonium Acetate.

[0346] 6) Transcription

[0347] Beads (Sample No.1) and precipitates (Samples No.2, No.3 andNo.4) were resuspended with components from the BioArray High YieldTranscription Kit (T7) (ENZO Diagnostics, N.Y.) and transcriptioncarried out in a 20 ul volume following the manufacturer's directionswith the addition of 5 uCi ³H-CTP , 20 Ci/mMol (Amersham PharmaciaBiotech, Piscataway, N.J.). In addition some reactions were carried outas described above, but T3 RNA polymerase from the BioArray High YieldTranscription Kit (T3) (ENZO Diagnostics, NY) was substituted. Reactionswere carried out for 120 minutes at 3720 C.

[0348] 7) Results 2^(nd) strand 3^(rd) strand Sample No. Primer PrimerRNA Polym CPM No. 1 T7-C9 — T7 12,392 No. 2 T7-C9 T3-T₂₅ T7 29,160 No. 2T7-C9 T3-T₂₅ T3 14,784 No. 3 C9 T7-T₂₅ T7 22,622 No. 4 C9 T3-T₂₅ T312,221

[0349] 8) Conclusions

[0350] This example demonstrated that a promoter can be introducedduring 3^(rd) strand synthesis to create functional production centers.This example also demonstrated that in addition to a T7 promoter, a T3promoter was also functional in the present method. This example alsodemonstrated that different production centers could be introduced intoeach end of a construct (Sample No.2) and both production centers werefunctional.

EXAMPLE 9 Multiple Rounds of 2^(nd) Strand Synthesis with ThermostablePolymerases

[0351] 1) Preparation of Beads, Binding of RNA to Beads and First StrandSynthesis were Carried out as Described in Example 8.

[0352] 2) Second Strand Synthesis and Recycling

[0353] Poly dG addition was carried out as described for sample No. 3 inExample 6 and the beads with tailed 3′ ends were used for 2^(nd) strandsynthesis under various conditions. 50 ul Reactions mixes were set up asfollows: Sample No.1 consisted of 1× Taq PCR Buffer (Epicentre, Madison,Wis.), 3 m M MgCl₂, 1× PCR Enhancer (Epicentre, Madison, Wis.), 0.4 mMdNTPs, 40 pMoles C9 primers and 5 units of Master Amp™ Taq DNAPolymerase (Epicentre, Madison, Wis.); Sample No.2 was the same assample No.1 except 100 pMoles of C9 primers were used; Sample No.3consisted of 1× Tth PCR Buffer (Epicentre, Madison, Wis.), 3 mM MgCl₂,1× PCR Enhancer (Epicentre, Madison, Wis.), 0.4 mM dNTPs, 40 pMoles C9primers and 5 units of Master Amp™ Tth DNA Polymerase (Epicentre,Madison, Wis.); Sample No.4 was the same as sample No.3 except 100pMoles of C9 primers were used Samples No.1 and No.3 went through onebinding/extension cycle while samples No.2 and No.4 went through 5 suchcycles. Each binding extension/extension cycle was carried out in athermocycler under the following conditions:

[0354] 2 minutes at 90° C.

[0355] 5 minutes at 4° C.

[0356] 5 minutes at 37° C.

[0357] 5 minutes at 50° C.

[0358] 15 minutes at 72° C.

[0359] At the end of each cycle, samples No.2 and No.4 were brieflyshaken to resuspend the beads. After the completion of either 1 or 5cycles, the mixtures were heated at 90° C. for 3 minutes and the aqueousportion collected after magnetic separation. Each sample was phenolextracted and ethanol precipitated as described previously in step 5 ofExample 8 for samples No.3 and No.4.

[0360] 3) Third Strand Synthesis

[0361] Pellets were resuspended in 26 ul of 10 mM Tris-HCl (pH 7.5) andT7-T₂₅ primers were added. For Samples No.1 and No.3, 40 pMoles ofT7-T₂₅ were added; for Samples No.2 and No.4, 400 pMoles of T7-T₂₅ wereadded. Third strand synthesis was then carried out by the addition ofMuLV, MuLV buffer and dNTPS as described in step 5 of Example 8.

[0362] 4) Transcription

[0363] Transcription was carried out as described previously without theaddition of radioactive precursors. Analysis of the reaction from eachsample was carried out by gel electophoreis as described previously andshown in FIG. 31.

[0364] 5) Conclusions

[0365] This example demonstrated that thermostable polymerases could beused for 2^(nd) strand synthesis in the methods described above. Thisexample also demonstrated that by increasing the amount of primers andthe number of cycles the amount of RNA copies derived from the originallibrary of nucleic acids was increased.

EXAMPLE 10 Levels of Transcription Derived from Sequential Rounds of2^(nd) strand Synthesis

[0366] 1) Preparation of Beads, Binding of RNA to Beads and First StrandSynthesis were Carried Out as Described in Example 8 Except the Amountof Analytes and Reagents for Each Reaction was Increased Two-Fold.Preparation of 1^(st) Strands for 2^(nd) Strand Synthesis was CarriedOut as Described Previously for Sample 3 in Example 6.

[0367] 2) Second Strand Synthesis

[0368] Second strand synthesis was carried out as described for SampleNo.3 in Example 8. Separation and isolation of the 2^(nd) strandproducts was carried out as described in Example 8 and set aside asSample No.1. Fresh reagents were then added to the beads and anotherround of 2^(nd) strand synthesis was carried out. The products of thissecond reaction were separated from the beads and designated SampleNo.2. The beads were then used once more for a third round of synthesis.The products of this reaction were set aside as Sample No.3.

[0369] 3) Third Strand Synthesis

[0370] Samples No.1, No.2 and No.3 were used as templates for 3^(rd)strand synthesis in individual reactions with the reagents and conditionpreviously described in Example 8. As mentioned above, the startingmaterial in the present example was twice the amount used in example 8and as such the amounts of all reagents were doubled for this reactionas well. For example, 80 pMoles of T7-T₂₅ primers were used.Purification of the products from each reaction was carried out asdescribed in Example 8.

[0371] 4) Transcription

[0372] Transcription reactions were carried out as with the BioArrayHigh Yield Transcription Kit (T7) (ENZO Diagnostics, N.Y.). The DNA wasused in a 20 ul final reaction volume which was incubated for 2 hours at37° C. Gel analysis was then used to evaluate the amount of synthesisthat was a result of each round of 2^(nd) strand synthesis describedabove. For purposes of contrast, various amounts of the transcriptionreaction (4 ul and 10 ul) were analyzed and in addition equvalentamounts of the DNA template that were not used in transcriptionreactions were also included. The results of this are shown in FIG. 32.

[0373] 5) Conclusion

[0374] This example demonstrated that the 2^(nd) strands made in eachround of 2^(nd) strand synthesis were substantially equal in theirability to be used to synthesize a library with functional productioncenters. FIG. 32 also shows the contrast between the amount oftranscript and the original DNA templates used for this synthesisthereby demonstrating the high levels of synthesis from each template.

EXAMPLE 11 Use of Reverse Transcriptases from Various Sources

[0375] Preparation of Beads, Binding of RNA to Beads and 1^(st) strandsynthesis were carried out as described in Example 6 except that ReverseTranscriptases from various sources were used for 1^(st) strandsynthesis reactions. 2^(nd) strand synthesis was carried out asdescribed in Example 6 for sample No.2 , i.e Terminal Transferaseaddition followed by binding and extension of T7-C9 primers. A list ofthe various Reverse Transcriptases and their sources is given below.

[0376] 1) Superscript II [RNaseH(−) MuLV] (Life Technologies, Rockville,Md.)

[0377] 2) RNase H (+) MuLV (Life Technologies, Rockville, Md.)

[0378] 3) RNase H (+) MuLV (New England Biolabs, Beverly, Mass.)

[0379] 4) Enhanced AMV (Sigma, St. Louis, Mo.)

[0380] 5) AMV (Life Technologies, Rockville, Md.)

[0381] 6) AMV (Sigma, St. Louis, Mo.)

[0382] 7) Omniscript (Qiagen

[0383] 8) Display THERMO-RT Display Systems Biotech,

[0384] 9) Powerscript [RNaseH(−) MuLV] (Clontech laboratories,

[0385] Each 2^(nd) stand synthesis was carried out in the bufferprovided by the manufacturer for each Reverse Transcriptase with theexception of the New England Biolabs version of RNase H (+) MuLV whichwas used in the buffer provided for the Life Technologies version ofRNase H (+) MuLV. Further processing and transcription reactions were aspreviously described in Example 6. The results of this experiment reshown in FIG. 33.

[0386] Conclusions

[0387] A variety of different Reverse Transcriptases ccould be used inconjunction with the methods of the present invention.

[0388] Many obvious variations will no doubt be suggested to those ofordinary skill in the art in light of the above detailed description andexamples of the present invention. All such variations are fullyembraced by the scope and spirit of the invention as more particularlydefined in the claims that now follow.

1 3 1 44 DNA Artificial Sequence Bacteriophage T7 Promoter with a C9 3′Tail 1 ggccagtgaa ttgtaatacg actcactata gggatccccc cccc 44 2 61 DNAArtificial Sequence Bacteriophage T7 Promoter with a T25 3′ Tail 2ggccagtgaa ttgtaatacg actcactata gggatctttt tttttttttt tttttttttt 60 t61 3 61 DNA Artificial Sequence Bacteriophage T3 Promoter with a T25 3′Tail 3 ctcaacgcca cctaattacc ctcactaaag ggagattttt tttttttttt tttttttttt60 t 61

What is claimed is:
 1. A composition of matter that comprises a libraryof analytes, said analytes being hybridized to an array of nucleicacids, said nucleic acids being fixed or immobilized to a solid support,wherein said analytes comprise an inherent universal detection target(UDT), and a universal detection element (UDE) attached to said UDTwherein said UDE generates a signal indicating the presence or quantityof said analytes, or said attachment of UDE to UDT.
 2. The compositionof claim 1, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 3. The composition of claim 1, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 4. The composition of claim 1, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 5. The composition of claim 4, wherein said analogs comprisePNA.
 6. The composition of claims 4 or 5, wherein said nucleic acids oranalogs are modified on any one of the sugar, phosphate or basemoieties.
 7. The composition of claim 1, wherein said solid support isporous or non-porous.
 8. The composition of claim 7, wherein said poroussolid support is selected from the group consisting of polyacrylamideand agarose.
 9. The composition of claim 7, wherein said non-poroussolid support comprises glass or plastic.
 10. The composition of claim1, wherein said solid support is transparent, translucent, opaque orreflective.
 11. The composition of claim 1, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.12. The composition of claim 11, wherein said nucleic acids areindirectly fixed or immobilized to said solid support by means of achemical linker or linkage arm.
 13. The composition of claim 1, whereinsaid inherent UDT is selected from the group consisting of 3′ polyAsegments, 5′ caps, secondary structures, consensus sequences and acombination of any of the foregoing.
 14. The composition of claim 13,wherein said consensus sequences is selected from the group consistingof signal sequences for polyA addition, splicing elements, multicopyrepeats and a combination of any of the foregoing.
 15. The compositionof claim 1, wherein said UDE is selected from the group consisting ofnucleic acids, nucleic acid analogs, polypeptides, polysaccharides,synthetic polymers and a combination of any of the foregoing.
 16. Thecomposition of claim 4, wherein said analogs comprise PNA.
 17. Thecomposition of claim 1, wherein said UDE generates a signal directly orindirectly.
 18. The composition of claim 17, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 19. The composition of claim 17, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 20. The composition ofclaim 19, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 21. A composition of matter that comprises alibrary of analytes, said analytes being hybridized to an array ofnucleic acids, said nucleic acids being fixed or immobilized to a solidsupport, wherein said analytes comprise a non-inherent universaldetection target (UDT) and a universal detection element (UDE)hybridized to said UDT, wherein said UDE generates a signal directly orindirectly to detect the presence or quantity of said analytes.
 22. Thecomposition of claim 21, wherein said library of analytes is derivedfrom a biological source selected from the group consisting of organs,tissues and cells.
 23. The composition of claim 21, wherein saidanalytes are selected from the group consisting of genomic DNA, episomalDNA, unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 24. The composition of claim 21, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 25. The composition of claim 24, wherein said analogs comprisePNA.
 26. The composition of claims 24 or 25, wherein said nucleic acidsor analogs are modified on any one of the sugar, phosphate or basemoieties.
 27. The composition of claim 21, wherein said solid support isporous or non-porous.
 28. The composition of claim 27, wherein saidporous solid support is selected from the group consisting ofpolyacrylamide and agarose.
 29. The composition of claim 27, whereinsaid non-porous solid support comprises glass or plastic.
 30. Thecomposition of claim 21, wherein said solid support is transparent,translucent, opaque or reflective.
 31. The composition of claim 21,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 32. The composition of claim 31,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 33. Thecomposition of claim 21, wherein said non-inherent universal detectiontarget (UDT) comprises homopolymeric sequences.
 34. The composition ofclaim of 21, wherein said non-inherent universal detection target (UDT)comprises heteropolymeric sequences.
 35. The composition of claim 21,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 36. The composition ofclaim 35, wherein said analogs comprise PNA.
 37. The composition ofclaim 21, wherein said UDE generates a signal directly or indirectly.38. The composition of claim 37, wherein said direct signal generationis selected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 39. The composition of claim 37, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 40. The composition ofclaim 39, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 41. A composition of matter that comprises alibrary of analytes, said analytes being hybridized to an array ofnucleic acids, said nucleic acids being fixed or immobilized to a solidsupport, wherein said hybridization between said analytes and saidnucleic acids generate a domain for complex formation, and saidcomposition further comprising a signaling entity complexed to saiddomain.
 42. The composition of claim 41, wherein said library ofanalytes is derived from a biological source selected from the groupconsisting of organs, tissues and cells.
 43. The composition of claim41, wherein said analytes are selected from the group consisting ofgenomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 44. The composition of claim 41,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 45. The composition of claim 44, whereinsaid analogs comprise PNA.
 46. The composition of claims 44 or 45,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 47. The composition of claim 41,wherein said solid support is porous or non-porous.
 48. The compositionof claim 47, wherein said porous solid support is selected from thegroup consisting of polyacrylamide and agarose.
 49. The composition ofclaim 47, wherein said non-porous solid support comprises glass orplastic.
 50. The composition of claim 41, wherein said solid support istransparent, translucent, opaque or reflective.
 51. The composition ofclaim 41, wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 52. The composition of claim 41,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 53. Thecomposition of claim 41, wherein said domain for complex formation isselected from the group consisting of DNA-DNA hybrids, DNA-RNA hybrids,RNA-RNA hybrids, DNA-PNA hybrids and RNA-PNA hybrids.
 54. Thecomposition of claim 41, wherein said signaling entity complexed to saiddomain is selected from the group consisting of proteins andintercalators.
 55. The composition of claim 54, wherein said proteinscomprise nucleic acid binding proteins which bind preferentially todouble-stranded nucleic acid.
 56. The composition of claim 55, whereinsaid nucleic acid binding proteins comprise antibodies.
 57. Thecomposition of claim 56, wherein said antibodies are specific fornucleic acid hybrids selected from the group consisting of DNA-DNAhybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids and RNA-PNAhybrids
 58. The composition of claim 54, wherein said intercalators areselected from the group consisting of ethidium bromide, diethidiumbromide, acridine orange and SYBR Green.
 59. The composition of claim41, wherein said proteins generate a signal directly or indirectly. 60.The composition of claim 59, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 61. The composition of claim 59, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 62. The composition ofclaim 61, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 63. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidscomplementary to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of said nucleic acidsof interest comprise at least one inherent universal detection target(UDT); and (iii) universal detection elements (UDE) which generates asignal directly or indirectly; b) hybridizing said library (ii) withsaid array of nucleic acids (i) to form hybrids if said nucleic acids ofinterest are present; c) contacting said UDEs with said UDTs to form acomplex bound to said array; d) detecting or quantifying said more thanone nucleic acid of interest by detecting or measuring the amount ofsignal generated from UDEs bound to said array.
 64. The process of claim63, wherein said nucleic acid array is selected from the groupconsisting of DNA, RNA and analogs thereof.
 65. The process of claim 64,wherein said analogs comprise PNA.
 66. The process of claims 64 or 65,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 67. The process of claim 63, whereinsaid solid support is porous or non-porous.
 68. The process of claim 67,wherein said porous solid support is selected from the group consistingof polyacrylamide and agarose.
 69. The process of claim 67, wherein saidnon-porous solid support comprises glass or plastic.
 70. The process ofclaim 63, wherein said solid support is transparent, translucent, opaqueor reflective.
 71. The process of claim 63, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.72. The process of claim 71, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 73. The process of claim 63, wherein said library ofanalytes is derived from a biological source selected from the groupconsisting of organs, tissues and cells.
 74. The process of claim 63,wherein said analytes are selected from the group consisting of genomicDNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and a combination ofany of the foregoing.
 75. The process of claim 63, wherein said inherentUDT is selected from the group consisting of 3′ polyA segments, 5′ caps,secondary structures, consensus sequences, and a combination of any ofthe foregoing.
 76. The process of claim 75, wherein said consensussequences is selected from the group consisting of signal sequences forpolyA addition, splicing elements, multicopy repeats, and a combinationof any of the foregoing.
 77. The process of claim 63, wherein said UDEis selected from the group consisting of nucleic acids, nucleic acidanalogs, polypeptides, polysaccharides, synthetic polymers and acombination of any of the foregoing.
 78. The process of claim 64,wherein said analogs comprise PNA.
 79. The process of claim 63, whereinsaid UDE generates a signal directly or indirectly.
 80. The process ofclaim 79, wherein said direct signal generation is selected from thegroup consisting of a fluorescent compound, a phosphorescent compound, achemiluminescent compound, a chelating compound, an electron densecompound, a magnetic compound, an intercalating compound, an energytransfer compound and a combination of any of the foregoing.
 81. Theprocess of claim 79, wherein said indirect signal generation is selectedfrom the group consisting of an antibody, an antigen, a hapten, areceptor, a hormone, a ligand, an enzyme and a combination of any of theforegoing.
 82. The process of claim 81, wherein said enzyme catalyzes areaction selected from the group consisting of a fluorogenic reaction, achromogenic reaction and a chemiluminescent reaction.
 83. The process ofclaim 63, comprising one or more washing steps.
 84. A process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of said nucleic acids of interest comprise at least one inherentuniversal detection target (UDT); and (iii) universal detection elements(UDE) which generates a signal directly or indirectly; b) contactingsaid UDEs with said UDTs in said library of nucleic acid analytes toform one or more complexes; c) hybridizing said library of nucleic acidanalytes with said array of nucleic acids (i) to form hybrids if saidnucleic acids of interest are present; d) detecting or quantifying saidmore than one nucleic acid of interest by detecting or measuring theamount of signal generated from UDEs bound to said array.
 85. Theprocess of claim 84, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 86. The process ofclaim 85, wherein said analogs comprise PNA.
 87. The process of claims85 or 86, wherein said nucleic acids or analogs are modified on any oneof the sugar, phosphate or base moieties.
 88. The process of claim 84,wherein said solid support is porous or non-porous.
 89. The process ofclaim 88, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 90. The process of claim 88,wherein said non-porous solid support comprises glass or plastic. 91.The process of claim 84, wherein said solid support is transparent,translucent, opaque or reflective.
 92. The process of claim 84, whereinsaid nucleic acids are directly or indirectly fixed or immobilized tosaid solid support.
 93. The process of claim 92, wherein said nucleicacids are indirectly fixed or immobilized to said solid support by meansof a chemical linker or linkage arm.
 94. The process of claim 84,wherein said library of analytes is derived from a biological sourceselected from the group consisting of organs, tissues and cells.
 95. Theprocess of claim 84, wherein said analytes are selected from the groupconsisting of genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA,snRNA and a combination of any of the foregoing.
 96. The process ofclaim 84, wherein said inherent UDT is selected from the groupconsisting of 3′ polyA segments, 5′ caps, secondary structures,consensus sequences, and a combination of any of the foregoing.
 97. Theprocess of claim 96, wherein said consensus sequences is selected fromthe group consisting of signal sequences for polyA addition, splicingelements, multicopy repeats, and a combination of any of the foregoing.98. The process of claim 84, wherein said UDE is selected from the groupconsisting of nucleic acids, nucleic acid analogs, polypeptides,polysaccharides, synthetic polymers and a combination of any of theforegoing.
 99. The process of claim 98, wherein said analogs comprisePNA.
 100. The process of claim 84, wherein said UDE generates a signaldirectly or indirectly.
 101. The process of claim 100, wherein saiddirect signal generation is selected from the group consisting of afluorescent compound, a phosphorescent compound, a chemiluminescentcompound, a chelating compound, an electron dense compound, a magneticcompound, an intercalating compound, an energy transfer compound and acombination of any of the foregoing.
 102. The process of claim 100,wherein said indirect signal generation is selected from the groupconsisting of an antibody, an antigen, a hapten, a receptor, a hormone,a ligand, an enzyme and a combination of any of the foregoing.
 103. Theprocess of claim 102, wherein said enzyme catalyzes a reaction selectedfrom the group consisting of a fluorogenic reaction, a chromogenicreaction and a chemiluminescent reaction.
 104. The process of claim 84,comprising one or more washing steps.
 105. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified, whereineach of said nucleic acids of interest comprise at least onenon-inherent universal detection target (UDT), wherein said non-inherentUDT is attached to said nucleic acid analytes; and (iii) universaldetection elements (UDE) which generate a signal directly or indirectly;b) hybridizing said library (ii) with said array of nucleic acids (i) toform hybrids if said nucleic acids of interest are present; c)contacting said UDEs with said UDTs to form a complex bound to saidarray; d) detecting or quantifying said more than one nucleic acid ofinterest by detecting or measuring the amount of signal generated fromUDEs bound to said array.
 106. The process of claim 105, wherein saidnucleic acid array is selected from the group consisting of DNA, RNA andanalogs thereof.
 107. The process of claim 106, wherein said analogscomprise PNA.
 108. The process of claims 106 or 107, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 109. The process of claim 105, wherein said solidsupport is porous or non-porous.
 110. The process of claim 109, whereinsaid porous solid support is selected from the group consisting ofpolyacrylamide and agarose.
 111. The process of claim 109, wherein saidnon-porous solid support comprises glass or plastic.
 112. The process ofclaim 105, wherein said solid support is transparent, translucent,opaque or reflective.
 113. The process of claim 105, wherein saidnucleic acids are directly or indirectly fixed or immobilized to saidsolid support.
 114. The process of claim 113, wherein said nucleic acidsare indirectly fixed or immobilized to said solid support by means of achemical linker or linkage arm.
 115. The process of claim 105, whereinsaid library of analytes is derived from a biological source selectedfrom the group consisting of organs, tissues and cells.
 116. The processof claim 105, wherein said analytes are selected from the groupconsisting of genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA,snRNA and a combination of any of the foregoing.
 117. The process ofclaim 105, wherein said non-inherent universal detection target (UDT)comprises homopolymeric sequences.
 118. The process of claim of 105,wherein said non-inherent universal detection target (UDT) comprisesheteropolymeric sequences.
 119. The process of claim 105, wherein saidUDE is selected from the group consisting of nucleic acids, nucleic acidanalogs and modified forms thereof.
 120. The process of claim 119,wherein said analogs comprise PNA.
 121. The process of claim 105,wherein said UDE generates a signal directly or indirectly.
 122. Theprocess of claim 121, wherein said direct signal generation is selectedfrom the group consisting of a fluorescent compound, a phosphorescentcompound, a chemiluminescent compound, a chelating compound, an electrondense compound, a magnetic compound, an intercalating compound, anenergy transfer compound and a combination of any of the foregoing. 123.The process of claim 121, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 124. The process of claim 123, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 125.The process of claim 105, comprising one or more washing steps.
 126. Aprocess for detecting or quantifying more than one nucleic acid ofinterest in a library comprising the steps of: a) providing: (i) anarray of fixed or immobilized nucleic acids complementary to saidnucleic acids of interest; (ii) a library of nucleic acid analytes whichmay contain the nucleic acids of interest sought to be detected orquantified, wherein each of said nucleic acids of interest comprise atleast one non-inherent universal detection target (UDT), wherein saidnon-inherent UDTs are attached to said nucleic acid analytes; and (iii)universal detection elements (UDE) which generate a signal directly orindirectly; b) contacting said UDEs with said UDTs in said library ofnucleic acid analytes to form one or more complexes; c) hybridizing saidlibrary (ii) with said array of nucleic acids (i) to form hybrids ifsaid nucleic acids of interest are present; d) detecting or quantifyingsaid more than one nucleic acid of interest by detecting or measuringthe amount of signal generated from UDEs bound to said array.
 127. Theprocess of claim 126, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 128. The processof claim 127, wherein said analogs comprise PNA.
 129. The process ofclaims 127 or 128, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 130. The process ofclaim 126, wherein said solid support is porous or non-porous.
 131. Theprocess of claim 130, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 132. The process ofclaim 130, wherein said non-porous solid support comprises glass orplastic.
 133. The process of claim 126, wherein said solid support istransparent, translucent, opaque or reflective.
 134. The process ofclaim 126, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 135. The process of claim 134,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 136. Theprocess of claim 126, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 137. The process of claim 126, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 138. The process of claim 126, wherein said non-inherentuniversal detection target (UDT) comprises homopolymeric sequences. 139.The process of claim of 126, wherein said non-inherent universaldetection target (UDT) comprises heteropolymeric sequences.
 140. Theprocess of claim 126, wherein said UDE is selected from the groupconsisting of nucleic acids, nucleic acid analogs and modified formsthereof.
 141. The process of claim 140, wherein said analogs comprisePNA.
 142. The process of claim 126, wherein said UDE generates a signaldirectly or indirectly.
 143. The process of claim 142, wherein saiddirect signal generation is selected from the group consisting of afluorescent compound, a phosphorescent compound, a chemiluminescentcompound, a chelating compound, an electron dense compound, a magneticcompound, an intercalating compound, an energy transfer compound and acombination of any of the foregoing.
 144. The process of claim 142,wherein said indirect signal generation is selected from the groupconsisting of an antibody, an antigen, a hapten, a receptor, a hormone,a ligand, an enzyme and a combination of any of the foregoing.
 145. Theprocess of claim 144, wherein said enzyme catalyzes a reaction selectedfrom the group consisting of a fluorogenic reaction, a chromogenicreaction and a chemiluminescent reaction.
 146. The process of claim 126,comprising one or more washing steps.
 147. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids complementary to said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)means for attaching one or more universal detection targets (UDT) to anucleic acid; (iv) universal detection elements (UDE) which generates asignal directly or indirectly; b) attaching said UDTs (iii) to saidlibrary of nucleic acid analytes (ii); c) hybridizing said library (ii)with said array of nucleic acids (i) to form hybrids if said nucleicacids of interest are present; d) contacting said UDEs with said UDTs toform a complex bound to said array; e) detecting or quantifying saidmore than one nucleic acid of interest by detecting or measuring theamount of signal generated from UDEs bound to said array.
 148. Theprocess of claim 147, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 149. The processof claim 148, wherein said analogs comprise PNA.
 150. The process ofclaims 148 or 149, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 151. The process ofclaim 147, wherein said solid support is porous or non-porous.
 152. Theprocess of claim 151, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 153. The process ofclaim 151, wherein said non-porous solid support comprises glass orplastic.
 154. The process of claim 147, wherein said solid support istransparent, translucent, opaque or reflective.
 155. The process ofclaim 147, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 156. The process of claim 155,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 157. Theprocess of claim 147, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 158. The process of claim 147, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 159. The process of claim 147, wherein said attaching meansadd homopolymeric sequences through an enzyme selected from the groupconsisting of poly A polymerase and terminal transferase.
 160. Theprocess of claim 147, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 161. The process of claim 147,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 162. The process ofclaim 161, wherein said analogs comprise PNA.
 163. The process of claim147, wherein said UDE generates a signal directly or indirectly. 164.The process of claim 163, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 165. The process of claim 163, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 166. The process ofclaim 165, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 167. The process of claim 147, comprising oneor more washing steps.
 168. A process for detecting or quantifying morethan one nucleic acid of interest in a library comprising the steps of:a) providing: (i) an array of fixed or immobilized nucleic acidscomplementary to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore universal detection targets (UDT) to a nucleic acid; (iv) universaldetection elements (UDE) which generates a signal directly orindirectly; b) attaching said UDTs (iii) to said library of nucleic acidanalytes (ii); c) contacting said UDEs with said UDTs in said library ofnucleic acid analytes to form one or more complexes; d) hybridizing saidlibrary (ii) with said array of nucleic acids (i) to form hybrids ifsaid nucleic acids of interest are present; e) detecting or quantifyingsaid more than one nucleic acid of interest by detecting or measuringthe amount of signal generated from UDEs bound to said array.
 169. Theprocess of claim 168, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 170. The processof claim 169, wherein said analogs comprise PNA.
 171. The process ofclaims 169 or 170, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 172. The process ofclaim 168, wherein said solid support is porous or non-porous.
 173. Theprocess of claim 172, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 174. The process ofclaim 172, wherein said non-porous solid support comprises glass orplastic.
 175. The process of claim 168, wherein said solid support istransparent, translucent, opaque or reflective.
 176. The process ofclaim 168, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 177. The process of claim 176,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 178. Theprocess of claim 168, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 179. The process of claim 168, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 180. The process of claim 168, wherein said attaching meansadd homopolymeric sequences through an enzyme selected from the groupconsisting of poly A polymerase and terminal transferase.
 181. Theprocess of claim 168, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 182. The process of claim 168,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 183. The process ofclaim 182, wherein said analogs comprise PNA.
 184. The process of claim168, wherein said UDE generates a signal directly or indirectly. 185.The process of claim 184, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 186. The process of claim 184, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 187. The process ofclaim 186, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 188. The process of claim 168, comprising oneor more washing steps.
 189. A process for detecting or quantifying morethan one nucleic acid of interest in a library comprising the steps of:a) providing: (i) an array of fixed or immobilized nucleic acidscomplementary to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; and (iii) universal detectionelements (UDEs) which bind to a domain formed by nucleic acid hybridsfor complex formation and generate a signal directly or indirectly; b)hybridizing said library (ii) with said array of nucleic acids (i) toform hybrids if said nucleic acids of interest are present, wherein saidformed hybrids generate a domain for complex formation; c) contactingsaid UDEs with said hybrids to form a complex bound to said array; d)detecting or quantifying said more than one nucleic acid of interest bydetecting or measuring the amount of signal generated from UDEs bound tosaid array.
 190. The process of claim 189, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 191. The process of claim 190, wherein said analogs comprisePNA.
 192. The process of claims 190 or 191, wherein said nucleic acidsor analogs are modified on any one of the sugar, phosphate or basemoieties.
 193. The process of claim 189, wherein said solid support isporous or non-porous.
 194. The process of claim 193, wherein said poroussolid support is selected from the group consisting of polyacrylamideand agarose.
 195. The process of claim 193, wherein said non-poroussolid support comprises glass or plastic.
 196. The process of claim 189,wherein said solid support is transparent, translucent, opaque orreflective.
 197. The process of claim 189, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.198. The process of claim 197, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 199. The process of claim 189, wherein said library ofanalytes is derived from a biological source selected from the groupconsisting of organs, tissues and cells.
 200. The process of claim 189,wherein said analytes are selected from the group consisting of genomicDNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and a combination ofany of the foregoing.
 201. The process of claim 189, wherein said domainfor complex formation is selected from the group consisting of DNA-DNAhybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids and RNA-PNAhybrids.
 202. The process of claim 189, wherein said signaling entitycomplexed to said domain is selected from the group consisting ofproteins and intercalators.
 203. The process of claim 202, wherein saidproteins comprise nucleic acid binding proteins which bindpreferentially to double-stranded nucleic acid.
 204. The process ofclaim 203, wherein said nucleic acid binding proteins compriseantibodies.
 205. The process of claim 204, wherein said antibodies arespecific for nucleic acid hybrids selected from the group consisting ofDNA-DNA hybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids andRNA-PNA hybrids.
 206. The process of claim 202, wherein saidintercalators are selected from the group consisting of ethidiumbromide, diethidium bromide, acridine orange and SYBR Green.
 207. Theprocess of claim 189, wherein said proteins generate a signal directlyor indirectly.
 208. The process of claim 207, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 209. The process of claim 207, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 210. The process ofclaim 209, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 211. The process of claim 189, furthercomprising one or more washing steps.
 212. A composition of mattercomprising a library of first nucleic acid analyte copies, said firstnucleic acid copies being hybridized to an array of nucleic acids, saidnucleic acids being fixed or immobilized to a solid support, whereinsaid first nucleic acid copies comprise an inherent universal detectiontarget (UDT) and a universal detection element (UDE) attached to saidUDT, wherein said UDE generates a signal directly or indirectly todetect the presence or quantity of said analytes.
 213. The compositionof claim 212, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 214. The composition of claim 212, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 215. The composition of claim 212, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 216. The composition of claim 215, wherein said analogscomprise PNA.
 217. The composition of claims 215 or 216, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 218. The composition of claim 212, wherein said solidsupport is porous or non-porous.
 219. The composition of claim 218,wherein said porous solid support is selected from the group consistingof polyacrylamide and agarose.
 220. The composition of claim 218,wherein said non-porous solid support comprises glass or plastic. 221.The composition of claim 212, wherein said solid support is transparent,translucent, opaque or reflective.
 222. The composition of claim 212,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 223. The composition of claim 222,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 224. Thecomposition of claim 212, wherein said inherent UDT is selected from thegroup consisting of poly T segments, secondary structures, consensussequences, and a combination of any of the foregoing.
 225. Thecomposition of claim 224, wherein said consensus sequences is selectedfrom the group consisting of signal sequences for polyA addition,splicing elements, multicopy repeats, and a combination of any of theforegoing.
 226. The composition of claim 212, wherein said UDE isselected from the group consisting of nucleic acids, nucleic acidanalogs, polypeptides, polysaccharides, synthetic polymers and acombination of any of the foregoing.
 227. The composition of claim 226,wherein said analogs comprise PNA.
 228. The composition of claim 212,wherein said UDE generates a signal directly or indirectly.
 229. Thecomposition of claim 228, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 230. The composition of claim 228, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 231. The compositionof claim 230, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 232. A composition of matter comprising alibrary of first nucleic acid copies, said first nucleic acid copiesbeing hybridized to an array of nucleic acids, said nucleic acids beingfixed or immobilized to a solid support, wherein said first nucleic acidcopies comprise one or more non-inherent universal detection targets(UDTs) and one or more universal detection elements (UDEs) attached tosaid UDTs, wherein said UDEs generate a signal directly or indirectly todetect the presence or quantity of said analytes, and wherein said UDTsare either: (i) at the 5′ ends of said first nucleic acid copies and notadjacent to an oligoT segment or sequence, or (ii) at the 3′ ends ofsaid first nucleic acid copies, or (iii) both (i) and (ii).
 233. Thecomposition of claim 232, wherein said library of analytes is derivedfrom a biological source selected from the group consisting of organs,tissues and cells.
 234. The composition of claim 232, wherein saidanalytes are selected from the group consisting of genomic DNA, episomalDNA, unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 235. The composition of claim 232, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 236. The composition of claim 235, wherein said analogscomprise PNA.
 237. The composition of claims 235 or 236, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 238. The composition of claim 232, wherein said solidsupport is porous or non-porous.
 239. The composition of claim 238,wherein said porous solid support is selected from the group consistingof polyacrylamide and agarose.
 240. The composition of claim 238,wherein said non-porous solid support comprises glass or plastic. 241.The composition of claim 232, wherein said solid support is transparent,translucent, opaque or reflective.
 242. The composition of claim 232,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 243. The composition of claim 242,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 244. Thecomposition of claim 232, wherein said non-inherent universal detectiontarget (UDT) comprises homopolymeric sequences.
 245. The composition ofclaim of 232, wherein said non-inherent universal detection target (UDT)comprises heteropolymeric sequences.
 246. The composition of claim 232,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs, polypeptides, polysaccharides, synthetic polymersand a combination of any of the foregoing.
 247. The composition of claim246, wherein said analogs comprise PNA.
 248. The composition of claim232, wherein said UDE generates a signal directly or indirectly. 249.The composition of claim 248, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 250. The composition of claim 248, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 251. The compositionof claim 250, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 252. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to said nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified, wherein each of saidnucleic acids of interest comprise at least one inherent universaldetection target (UDT); (iii) universal detection elements (UDE) whichgenerate a signal directly or indirectly; and (iv) polymerizing meansfor synthesizing nucleic acid copies of said nucleic acids of analytes;b) synthesizing one or more first nucleic acid copies which arecomplementary to all or part of said nucleic acid analytes andsynthesizing sequences which are complementary to all or part of saidUDT to form a complementary UDT; c) hybridizing said first nucleic acidcopies with said array of nucleic acids (i) to form hybrids if saidnucleic acids of interest are present; d) contacting said UDEs with saidcomplementary UDTs of said first nucleic acid copies to form a complexbound to said array; e) detecting or quantifying said more than onenucleic acid of interest by detecting or measuring the amount of signalgenerated from UDEs bound to said array.
 253. The process of claim 252,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 254. The process of claim 253, whereinsaid analogs comprise PNA.
 255. The process of claims 253 or 254,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 256. The process of claim 252,wherein said solid support is porous or non-porous.
 257. The process ofclaim 256, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 258. The process of claim 256,wherein said non-porous solid support comprises glass or plastic. 259.The process of claim 252, wherein said solid support is transparent,translucent, opaque or reflective.
 260. The process of claim 252,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 261. The process of claim 260,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 262. Theprocess of claim 252, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 263. The process of claim 252, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 264. The process of claim 252, wherein said inherent UDT isselected from the group consisting of poly T segments, secondarystructures, consensus sequences, and a combination of any of theforegoing.
 265. The process of claim 264, wherein said consensussequences is selected from the group consisting of signal sequences forpolyA addition, splicing elements, multicopy repeats, and a combinationof any of the foregoing.
 266. The process of claim 252, wherein said UDEis selected from the group consisting of nucleic acids, nucleic acidanalogs, polypeptides, polysaccharides, synthetic polymers and acombination of any of the foregoing.
 267. The process of claim 266,wherein said analogs comprise PNA.
 268. The process of claim 252,wherein said UDE generates a signal directly or indirectly.
 269. Theprocess of claim 268, wherein said direct signal generation is selectedfrom the group consisting of a fluorescent compound, a phosphorescentcompound, a chemiluminescent compound, a chelating compound, an electrondense compound, a magnetic compound, an intercalating compound, anenergy transfer compound and a combination of any of the foregoing. 270.The process of claim 268, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 271. The process of claim 270, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 272.The process of claim 252, wherein said polymerizing means are selectedfrom the group consisting of E. coli DNA Pol I, Klenow fragment of E.coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNApolymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.
 273. A process for detecting or quantifying more thanone nucleic acid of interest in a library comprising the steps of: a)providing: (i) an array of fixed or immobilized nucleic acids identicalin part or whole to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified, wherein each of said nucleic acidsof interest comprise at least one inherent universal detection target(UDT); (iii) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (iv) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes; b) synthesizing oneor more first nucleic acid copies of said nucleic acid analytes; c)contacting said UDEs with said UDTs in said first nucleic acid copies toform one or more complexes; d) hybridizing said first nucleic acidcopies with said array of nucleic acids (i) to form hybrids if saidnucleic acids of interest are present; and e) detecting or quantifyingsaid more than one nucleic acid of interest by detecting or measuringthe amount of signal generated from UDEs bound to said array.
 274. Theprocess of claim 273, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 275. The processof claim 274, wherein said analogs comprise PNA.
 276. The process ofclaims 274 or 275, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 277. The process ofclaim 273, wherein said solid support is porous or non-porous.
 278. Theprocess of claim 277, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 279. The process ofclaim 277, wherein said non-porous solid support comprises glass orplastic.
 280. The process of claim 273, wherein said solid support istransparent, translucent, opaque or reflective.
 281. The process ofclaim 273, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 282. The process of claim 281,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 283. Theprocess of claim 273, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 284. The process of claim 273, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 285. The process of claim 273, wherein said inherent UDT isselected from the group consisting of poly T segments, secondarystructures, consensus sequences, and a combination of any of theforegoing.
 286. The process of claim 285, wherein said consensussequences is selected from the group consisting of signal sequences forpolyA addition, splicing elements, multicopy repeats, and a combinationof any of the foregoing.
 287. The process of claim 273, wherein said UDEis selected from the group consisting of nucleic acids, nucleic acidanalogs, polypeptides, polysaccharides, synthetic polymers and acombination of any of the foregoing.
 288. The process of claim 287,wherein said analogs comprise PNA.
 289. The process of claim 273,wherein said UDE generates a signal directly or indirectly.
 290. Theprocess of claim 289, wherein said direct signal generation is selectedfrom the group consisting of a fluorescent compound, a phosphorescentcompound, a chemiluminescent compound, a chelating compound, an electrondense compound, a magnetic compound, an intercalating compound, anenergy transfer compound and a combination of any of the foregoing. 291.The process of claim 289, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 292. The process of claim 291, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 293.The process of claim 283, wherein said polymerizing means are selectedfrom the group consisting of E. coli DNA Pol I, Klenow fragment of E.coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNApolymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.
 294. A process for detecting or quantifying more thanone nucleic acid of interest in a library comprising the steps of: a)providing: (i) an array of fixed or immobilized nucleic acids identicalin part or whole to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) means for attaching one ormore non-inherent universal detection targets (UDT) to a nucleic acid;(iv) universal detection elements (UDE) which generate a signal directlyor indirectly; and (v) polymerizing means for synthesizing nucleic acidcopies of said nucleic acid analytes; b) attaching said non-inherentUDTs to either the 3′ ends of said nucleic acid analytes, the 5′ ends ofsaid first nucleic acid analytes, or both said 3′ ends and said 5′ endsof said nucleic acid analytes; c) synthesizing one or more first nucleicacid copies of said nucleic acid analytes; d) hybridizing said firstnucleic acid copies with said array of nucleic acids (i) to form hybridsif said nucleic acids of interest are present; e) contacting said UDEswith said UDTs of said first nucleic acid copies to form a complex boundto said array; and f) detecting or quantifying said more than onenucleic acid of interest by detecting or measuring the amount of signalgenerated from UDEs bound to said array.
 295. The process of claim 294,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 296. The process of claim 295, whereinsaid analogs comprise PNA.
 297. The process of claims 295 or 296,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 298. The process of claim 294,wherein said solid support is porous or non-porous.
 299. The process ofclaim 298, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 300. The process of claim 298,wherein said non-porous solid support comprises glass or plastic. 301.The process of claim 294, wherein said solid support is transparent,translucent, opaque or reflective.
 302. The process of claim 294,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 303. The process of claim 302,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 304. Theprocess of claim 294, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 305. The process of claim 294, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 306. The process of claim 294, wherein said attaching meansadd homopolymeric sequences through an enzyme selected from the groupconsisting of poly A polymerase and terminal transferase.
 307. Theprocess of claim 294, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 308. The process of claim 294,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 309. The process ofclaim 308, wherein said analogs comprise PNA.
 310. The process of claim294, wherein said UDE generates a signal directly or indirectly. 311.The process of claim 310, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 312. The process of claim 310, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 313. The process ofclaim 312, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 314. The process of claim 294, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 315. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to said nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) means for attachingone or more non-inherent universal detection targets (UDT) to a nucleicacid; (iv) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (v) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes; b) attaching saidnon-inherent UDTs to either the 3′ ends of said nucleic acid analytes,the 5′ ends of said first nucleic acid analytes, or both said 3′ endsand said 5′ ends of said nucleic acid analytes; c) synthesizing one ormore first nucleic acid copies of said nucleic acid analytes; d)contacting said UDEs with said UDTs of said first nucleic acid copies toform complexes; e) hybridizing said first nucleic acid copies with saidarray of nucleic acids (i) to form hybrids if said nucleic acids ofinterest are present; f) detecting or quantifying said more than onenucleic acid of interest by detecting or measuring the amount of signalgenerated from UDEs bound to said array.
 316. The process of claim 315,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 317. The process of claim 316, whereinsaid analogs comprise PNA.
 318. The process of claims 316 or 317,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 319. The process of claim 315,wherein said solid support is porous or non-porous.
 320. The process ofclaim 319, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 321. The process of claim 319,wherein said non-porous solid support comprises glass or plastic. 322.The process of claim 315, wherein said solid support is transparent,translucent, opaque or reflective.
 323. The process of claim 315,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 324. The process of claim 323,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 325. Theprocess of claim 315, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 326. The process of claim 315, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 327. The process of claim 315, wherein said attaching meansadd homopolymeric sequences through an enzyme selected from the groupconsisting of poly A polymerase and terminal transferase.
 328. Theprocess of claim 315, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 329. The process of claim 315,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 330. The process ofclaim 329, wherein said analogs comprise PNA.
 331. The process of claim315, wherein said UDE generates a signal directly or indirectly. 332.The process of claim 331, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 333. The process of claim 331, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 334. The process ofclaim 333, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 335. The process of claim 315, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 336. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to said nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) means for attachingone or more non-inherent universal detection targets (UDT) to a nucleicacid; (iv) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (v) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes; b) synthesizing oneor more first nucleic acid copies of said nucleic acid analytes; c)attaching said non-inherent UDTs to either the 3′ ends of said firstnucleic acid copies, the 5′ ends of said first nucleic acid copies, orboth said 3′ ends and said 5′ ends of said first nucleic acid copies; d)hybridizing said first nucleic acid copies with said array of nucleicacids (i) to form hybrids if said nucleic acids of interest are present;e) contacting said UDEs with said UDTs of said first nucleic acid copiesto form a complex bound to said array; f) detecting or quantifying saidmore than one nucleic acid of interest by detecting or measuring theamount of signal generated from UDEs bound to said array.
 337. Theprocess of claim 336, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 338. The processof claim 337, wherein said analogs comprise PNA.
 339. The process ofclaims 337 or 338, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 340. The process ofclaim 336, wherein said solid support is porous or non-porous.
 341. Theprocess of claim 340, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 342. The process ofclaim 340, wherein said non-porous solid support comprises glass orplastic.
 343. The process of claim 336, wherein said solid support istransparent, translucent, opaque or reflective.
 344. The process ofclaim 336, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 345. The process of claim 344,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 346. Theprocess of claim 336, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 347. The process of claim 336, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 348. The process of claim 336, wherein said attaching meansadd homopolymeric sequences through terminal transferase.
 349. Theprocess of claim 336, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 350. The process of claim 336,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 351. The process ofclaim 350, wherein said analogs comprise PNA.
 352. The process of claim336, wherein said UDE generates a signal directly or indirectly. 353.The process of claim 352, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 354. The process of claim 352, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 355. The process ofclaim 354, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 356. The process of claim 336, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 357. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to said nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; (iii) means for attachingone or more non-inherent universal detection targets (UDT) to a nucleicacid; (iv) universal detection elements (UDE) which generate a signaldirectly or indirectly; and (v) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes; b) synthesizing oneor more first nucleic acid copies of said nucleic acid analytes; c)attaching said non-inherent UDTs to either the 3′ ends of said firstnucleic acid copies, the 5′ ends of said first nucleic acid copies, orboth said 3′ ends and said 5′ ends of said first nucleic acid copies; d)contacting said UDEs with said UDTs of said first nucleic acid copies toform a complex; e) hybridizing said first nucleic acid copies with saidarray of nucleic acids (i) to form hybrids if said nucleic acids ofinterest are present; and f) detecting or quantifying said more than onenucleic acid of interest by detecting or measuring the amount of signalgenerated from UDEs bound to said array.
 358. The process of claim 357,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 359. The process of claim 358, whereinsaid analogs comprise PNA.
 360. The process of claims 358 or 359,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 361. The process of claim 357,wherein said solid support is porous or non-porous.
 362. The process ofclaim 361, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 363. The process of claim 361,wherein said non-porous solid support comprises glass or plastic. 364.The process of claim 357, wherein said solid support is transparent,translucent, opaque or reflective.
 365. The process of claim 357,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 366. The process of claim 365,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 367. Theprocess of claim 357, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 368. The process of claim 357, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 369. The process of claim 357, wherein said attaching meansadd homopolymeric sequences through terminal transferase.
 370. Theprocess of claim 357, wherein said attaching means add homopolymeric orheteropolymeric sequences through an enzyme selected from the groupconsisting of DNA ligase and RNA ligase.
 371. The process of claim 357,wherein said UDE is selected from the group consisting of nucleic acids,nucleic acid analogs and modified forms thereof.
 372. The process ofclaim 371, wherein said analogs comprise PNA.
 373. The process of claim357, wherein said UDE generates a signal directly or indirectly. 374.The process of claim 373, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 375. The process of claim 373, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 376. The process ofclaim 375, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 377. The process of claim 357, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 378. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of fixed or immobilized nucleic acidscomplementary to said nucleic acids of interest; (ii) a library ofnucleic acid analytes which may contain the nucleic acids of interestsought to be detected or quantified; (iii) universal detection elements(UDEs) which bind to a domain for complex formation formed by nucleicacid hybrids and generate a signal directly or indirectly; and (iv)polymerizing means for synthesizing nucleic acid copies of said nucleicacid analytes; b) synthesizing one or more nucleic acid copies of saidnucleic acid analytes; c) hybridizing said first nucleic acid copieswith said array of nucleic acids (i) to form hybrids if said nucleicacids of interest are present, wherein said formed hybrids generate adomain for complex formation; d) contacting said UDEs with said hybridsto form a complex bound to said array; and e) detecting or quantifyingsaid more than one nucleic acid of interest by detecting or measuringthe amount of signal generated from UDEs bound to said array.
 379. Theprocess of claim 378, wherein said nucleic acid array is selected fromthe group consisting of DNA, RNA and analogs thereof.
 380. The processof claim 379, wherein said analogs comprise PNA.
 381. The process ofclaims 379 or 380, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 382. The process ofclaim 378, wherein said solid support is porous or non-porous.
 383. Theprocess of claim 382, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 384. The process ofclaim 382, wherein said non-porous solid support comprises glass orplastic.
 385. The process of claim 378, wherein said solid support istransparent, translucent, opaque or reflective.
 386. The process ofclaim 378, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 387. The process of claim 386,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 388. Theprocess of claim 378, wherein said library of analytes is derived from abiological source selected from the group consisting of organs, tissuesand cells.
 389. The process of claim 378, wherein said analytes areselected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 390. The process of claim 378, wherein said domain forcomplex formation is selected from the group consisting of DNA-DNAhybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids and RNA-PNAhybrids.
 391. The process of claim 378, wherein said signaling entitycomplexed to said domain is selected from the group consisting ofproteins and intercalators.
 392. The process of claim 391, wherein saidproteins comprise nucleic acid binding proteins which bindpreferentially to double-stranded nucleic acid.
 393. The process ofclaim 392, wherein said nucleic acid binding proteins compriseantibodies.
 394. The process of claim 393, wherein said antibodies arespecific for nucleic acid hybrids selected from the group consisting ofDNA-DNA hybrids, DNA-RNA hybrids, RNA-RNA hybrids, DNA-PNA hybrids andRNA-PNA hybrids
 395. The process of claim 391, wherein saidintercalators are selected from the group consisting of ethidiumbromide, diethidium bromide, acridine orange and SYBR Green.
 396. Theprocess of claim 391, wherein said protein generates a signal directlyor indirectly.
 397. The process of claim 396, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 398. The process of claim 396, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 399. The process ofclaim 398, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 400. The process of claim 378, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 401. A composition of matter comprising alibrary of double-stranded nucleic acids substantially incapable of invivo replication and free of non-inherent homopolymeric sequences, saidnucleic acids comprising sequences complementary or identical in part orwhole to inherent sequences of a library obtained from a sample, whereinsaid double-stranded nucleic acids comprise at least one inherentuniversal detection target (UDT) proximate to one end of said doublestrand and at least one non-inherent production center proximate to theother end of said double strand.
 402. The composition of claim 401,wherein said sample comprises a biological a source selected from thegroup consisting of organs, tissues and cells.
 403. The composition ofclaim 401, wherein said library of nucleic acids are derived from thegroup consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.
 404. Thecomposition of claim 401, wherein said inherent UDT is selected from thegroup consisting of 3′ polyA segments, consensus sequences, or both.405. The composition of claim 404, wherein said consensus sequences isselected from the group consisting of signal sequences for poly Aaddition, splicing elements, multicopy repeats, and a combination of anyof the foregoing.
 406. The composition of claim 401, wherein saidproduction center is selected from the group consisting of primerbinding sites, RNA promoters, or a combination of both.
 407. Thecomposition of claim 406, wherein said RNA promoters comprise phagepromoters.
 408. The composition of claim 407, wherein said phagepromoters are selected from the group consisting of T3, T7 and SP6. 409.A composition of matter comprising a library of double-stranded nucleicacids substantially incapable of in vivo replication, said nucleic acidscomprising sequences complementary or identical in part or whole toinherent sequences of a library obtained from a sample, wherein saiddouble-stranded nucleic acids comprise at least four (4) non-inherentnucleotides proximate to one end of said double strand and anon-inherent production center proximate to the other end of said doublestrand.
 410. The composition of claim 409, wherein said sample comprisesa biological source selected from the group consisting of organs,tissues and cells.
 411. The composition of claim 409, wherein saidlibrary of nucleic acids are derived from the group consisting ofgenomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 412. The composition of claim 409,further comprising one or more inherent UDTs selected from the groupconsisting of 3′ polyA segments, consensus sequences, or both.
 413. Thecomposition of claim 412, wherein said consensus sequences is selectedfrom the group consisting of signal sequences for polyA addition,splicing elements, multicopy repeats, and a combination of any of theforegoing.
 414. The composition of claim 409, wherein said at least four(4) non-inherent nucleotides are homopolymeric.
 415. The composition ofclaim 409, wherein said non-inherent production center is selected fromthe group consisting of primer binding sites, RNA promoters, or acombination of both.
 416. The composition of claim 415, wherein said RNApromoters comprise phage promoters.
 417. The composition of claim 416,wherein said phage promoters are selected from the group consisting ofT3, T7 and SP6.
 418. A composition of matter comprising a library ofdouble-stranded nucleic acids fixed to a solid support, said nucleicacids comprising sequences complementary or identical in part or wholeto inherent sequences of a library obtained from a sample and saidnucleic acids further comprising at least one first sequence segment ofnon-inherent nucleotides proximate to one end of said double strand andat least one second sequence segment proximate to the other end of saiddouble strand, said second sequence segment comprising at least oneproduction center.
 419. The composition of claim 418, wherein said solidsupport comprises beads.
 420. The composition of claim 419, wherein saidbeads are magnetic.
 421. The composition of claim 418, wherein saidsample comprises a biological source selected from the group consistingof organs, tissues and cells.
 422. The composition of claim 418, whereinsaid library of nucleic acids are derived from the group consisting ofgenomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 423. The composition of claim 418,further comprising one or more inherent UDTs selected from the groupconsisting of 3′ poly A segments, consensus sequences, or both.
 424. Thecomposition of claim 423, wherein said consensus sequences is selectedfrom the group consisting of signal sequences for poly A addition,splicing elements, multicopy repeats, and a combination of any of theforegoing.
 425. The composition of claim 418, wherein said non-inherentproduction center is selected from the group consisting of primerbinding sites, RNA promoters, or a combination of both.
 426. Thecomposition of claim 425, wherein said RNA promoters comprise phagepromoters.
 427. The composition of claim 426, wherein said phagepromoters are selected from the group consisting of T3, T7 and SP6. 428.A composition of matter comprising a library of double-stranded nucleicacids attached to a solid support, said nucleic acids comprisingsequences complementary or identical in part or whole to inherentsequences of a library obtained from a sample, wherein saiddouble-stranded nucleic acids comprise at least one inherent universaldetection target (UDT) proximate to one end of said double strand and atleast one non-inherent production center proximate to the other end ofsaid double strand.
 429. The composition of claim 428, wherein saidsolid support comprises beads.
 430. The composition of claim 429,wherein said beads are magnetic.
 431. The composition of claim 428,wherein said sample comprises a biological source selected from thegroup consisting of organs, tissues and cells.
 432. The composition ofclaim 428, wherein said library of nucleic acids are derived from thegroup consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.
 433. Thecomposition of claim 428, wherein said inherent UDT is selected from thegroup consisting of 3′ polyA segments, consensus sequences, or both.434. The composition of claim 433, wherein said consensus sequences isselected from the group consisting of signal sequences for polyAaddition, splicing elements, multicopy repeats, and a combination of anyof the foregoing.
 435. The composition of claim 428, wherein saidproduction center is selected from the group consisting of primerbinding sites, RNA promoters, or a combination of both.
 436. Thecomposition of claim 435, wherein said RNA promoters comprise phagepromoters.
 437. The composition of claim 436, wherein said phagepromoters are selected from the group consisting of T3, T7 and SP6. 438.A process for detecting or quantifying more than one nucleic acid ofinterest in a library comprising the steps of: a) providing: (i) anarray of fixed or immobilized nucleic acids identical or complementaryin part or whole to sequences of said nucleic acids of interest; (ii) alibrary of nucleic acid analytes which may contain the nucleic acids ofinterest sought to be detected or quantified; and (iii) polymerizingmeans for synthesizing nucleic acid copies of said nucleic acidanalytes, said polymerizing means comprising a first set of primers anda second set of primers, wherein said second set of primers comprises atleast two segments, the first segment at the 3′ end comprising randomsequences, and the second segment comprising at least one productioncenter; (iv) means for synthesizing nucleic acid copies under isothermalor isostatic conditions; b) contacting said library of nucleic acidanalytes with said first set of primers to form more than one firstbound entity; c) extending said bound first set of primers by means oftemplate sequences provided by said nucleic acid analytes to form firstcopies of said analytes; d) contacting said extended first copies withsaid second set of primers to form more than one second bound entity; e)extending said bound second set of primers by means of templatesequences provided by said extended first copies to form more than onecomplex comprising extended first copies and extended second set ofprimers; f) synthesizing from a production center in said second set ofprimers in said complexes one or more nucleic acid copies underisothermal or isostatic conditions; g) hybridizing said nucleic acidcopies formed in step f) to said array of nucleic acids provided in stepa) (i); and h) detecting or quantifying any of said hybridized copiesobtained in step g).
 439. The process of claim 438, wherein said nucleicacid array comprises members selected from the group consisting of DNA,RNA and analogs thereof.
 440. The process of claim 439, wherein saidanalogs comprise PNA.
 441. The process of claims 439 or 440, whereinsaid nucleic acids or analogs are modified on any one of the sugar,phosphate or base moieties.
 442. The process of claim 438, wherein saidnucleic acid array is fixed or immobilized to a solid support.
 443. Theprocess of claim 442, wherein said solid support is porous ornon-porous.
 444. The process of claim 443, wherein said porous solidsupport is selected from the group consisting of polyacrylamide andagarose.
 445. The process of claim 443, wherein said non-porous solidsupport comprises glass or plastic.
 446. The process of claim 442,wherein said solid support is transparent, translucent, opaque orreflective.
 447. The process of claim 442, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.448. The process of claim 447, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 449. The process of claim 438, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 450. The process ofclaim 438, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 451. Theprocess of claim 438, wherein said first set of primers arecomplementary to inherent UDTs.
 452. The process of claim 438, whereinsaid inherent UDT is selected from the group consisting of 3′ poly Asegments, consensus sequences, and a combination of both.
 453. Theprocess of claim 452, wherein said consensus sequences is selected fromthe group consisting of signal sequences for poly A addition, splicingelements, multicopy repeats, and a combination of any of the foregoing.454. The process of claim 438, wherein said production center isselected from the group consisting of primer binding sites, RNApromoters, or a combination of both.
 455. The process of claim 454,wherein said RNA promoters comprise phage promoters.
 456. The process ofclaim 455, wherein said phage promoters are selected from the groupconsisting of T3, T7 and SP6.
 457. The process of claim 438, whereinsaid hybridized nucleic acid copies further comprise one or moresignaling entities attached or incorporated thereto.
 458. The process ofclaim 457, wherein said signaling entities generate a signal directly orindirectly.
 459. The process of claim 458, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 460. The process of claim 458, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 461. The process ofclaim 460, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 462. The process of claim 438, furthercomprising the step of separating the first copies obtained from step c)from their templates and repeating step b).
 463. The process of claim438, further comprising the step of separating the extended second setof primers obtained from step f) from their templates and repeating stepe).
 464. The process of claim 438, wherein step g) is carried outrepeatedly.
 465. The process of claim 438, wherein said polymerizingmeans are selected from the group consisting of E. coli DNA Pol I,Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 466. The process of claim 438, wherein saidmeans for synthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 467. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers and a second set of primers,wherein said first set of primers comprise at least one productioncenter; and (iv) means for synthesizing nucleic acid copies underisothermal or isostatic conditions; b) contacting said library ofnucleic acid analytes with said first set of primers to form more thanone first bound entity; c) extending said bound first set of primers bymeans of template sequences provided by said nucleic acid analytes toform first copies of said analytes; d) extending said first copies bymeans of at least four (4) or more non-inherent homopolymericnucleotides; e) contacting said extended first copies with said secondset of primers to form more than one second bound entity; f) extendingsaid bound second set of primers by means of template sequences providedby said extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; g)synthesizing from a production center in said second set of primers insaid complexes one or more nucleic acid copies under isothermal orisostatic conditions; h) hybridizing said nucleic acid copies formed instep g) to said array of nucleic acids provided in step a) (i); and i)detecting or quantifying any of said hybridized copies obtained in steph).
 468. The process of claim 467, wherein said nucleic acid arraycomprises members selected from the group consisting of DNA, RNA andanalogs thereof.
 469. The process of claim 468, wherein said analogscomprise PNA.
 470. The process of claims 468 or 469, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 471. The process of claim 467, wherein said nucleicacid array is fixed or immobilized to a solid support.
 472. The processof claim 471, wherein said solid support is porous or non-porous. 473.The process of claim 472, wherein said porous solid support is selectedfrom the group consisting of polyacrylamide and agarose.
 474. Theprocess of claim 472, wherein said non-porous solid support comprisesglass or plastic.
 475. The process of claim 471, wherein said solidsupport is transparent, translucent, opaque or reflective.
 476. Theprocess of claim 471, wherein said nucleic acids are directly orindirectly fixed or immobilized to said solid support.
 477. The processof claim 471, wherein said nucleic acids are indirectly fixed orimmobilized to said solid support by means of a chemical linker orlinkage arm.
 478. The process of claim 467, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 479. The process ofclaim 467, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 480. Theprocess of claim 467, wherein said first set of primers further compriseone or more sequences complementary to inherent universal detectiontargets (UDTs).
 481. The process of claim 467, wherein said inherent UDTis selected from the group consisting of 3′ poly A segments, consensussequences, and a combination of both.
 482. The process of claim 481,wherein said consensus sequences is selected from the group consistingof signal sequences for poly A addition, splicing elements, multicopyrepeats, and a combination of any of the foregoing.
 483. The process ofclaim 467, wherein said production center is selected from the groupconsisting of primer binding sites, RNA promoters, or a combination ofboth.
 484. The process of claim 483, wherein said RNA promoters comprisephage promoters.
 485. The process of claim 484, wherein said phagepromoters are selected from the group consisting of T3, T7 and SP6. 486.The process of claim 467, wherein said extending step d), the four ormore non-inherent homopolymeric nucleotides are added by terminaltransferase.
 487. The process of claim 467, wherein said hybridizednucleic acid copies further comprise one or more signaling entitiesattached or incorporated thereto.
 488. The process of claim 487, whereinsaid signaling entities generate a signal directly or indirectly. 489.The process of claim 488, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 490. The process of claim 489, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 491. The process ofclaim 490, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 492. The process of claim 467, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 493. The process of claim 467, wherein saidmeans for synthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 494. The process of claim 467,further comprising the step of separating the first copies obtained fromstep c) from their templates and repeating step b).
 495. The process ofclaim 467, further comprising the step of separating the extended secondset of primers obtained from step f) from their templates and repeatingstep e).
 496. The process of claim 467, wherein step g) is carried outrepeatedly.
 497. A process for detecting or quantifying more than onenucleic acid of interest in a library comprising the steps of: a)providing: (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of said nucleicacid analytes, said polymerizing means comprising a first set of primersand a second set of primers, wherein said first set comprises at leastone production center; (iv) a set of oligonucleotides or polynucleotidescomplementary to at least one segment or sequence of said second set ofprimers; and (v) means for ligating said set of oligonucleotides orpolynucleotides (iv); b) contacting said library of nucleic acidanalytes with said first set of primers to form more than one firstbound entity; c) extending said bound first set of primers by means oftemplate sequences provided by said nucleic acid analytes to form firstcopies of said analytes; d) ligating said set of oligonucleotides orpolynucleotides a) (iv) to the 3′ end of said first copies formed instep c) to form more than one ligated product; e) contacting saidligated product with said second set of primers to form more than onesecond bound entity; f) extending said bound second set of primers bymeans of template sequences provided by said ligated products formed instep d) to form more than one complex comprising said ligated productsand said extended second set of primers; g) synthesizing from aproduction center in said second set of primers in said complexes one ormore nucleic acid copies under isothermal or isostatic conditions; h)hybridizing said nucleic acid copies formed in step g) to said array ofnucleic acids provided in step a) (i); and i) detecting or quantifyingany of said hybridized copies obtained in step h).
 498. The process ofclaim 497, wherein said nucleic acid array comprises members selectedfrom the group consisting of DNA, RNA and analogs thereof.
 499. Theprocess of claim 498, wherein said analogs comprise PNA.
 500. Theprocess of claims 498 or 499, wherein said nucleic acids or analogs aremodified on any one of the sugar, phosphate or base moieties.
 501. Theprocess of claim 497, wherein said nucleic acid array is fixed orimmobilized to a solid support.
 502. The process of claim 501, whereinsaid solid support is porous or non-porous.
 503. The process of claim502, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 504. The process of claim 502,wherein said non-porous solid support comprises glass or plastic. 505.The process of claim 501, wherein said solid support is transparent,translucent, opaque or reflective.
 506. The process of claim 501,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 507. The process of claim 506,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 508. Theprocess of claim 497, wherein said library of nucleic acid analytes isderived from a biological source selected from the group consisting oforgans, tissues and cells.
 509. The process of claim 497, wherein saidlibrary of nucleic acids analytes are derived from the group consistingof genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 510. The process of claim 497,wherein said first set of primers are complementary to inherentuniversal detection targets (UDTs).
 511. The process of claim 497,wherein said inherent UDTs are selected from the group consisting of 3′poly A segments, consensus sequences, and a combination of both. 512.The process of claim 511, wherein said consensus sequences is selectedfrom the group consisting of signal sequences for poly A addition,splicing elements, multicopy repeats, and a combination of any of theforegoing.
 513. The process of claim 497, wherein said production centeris selected from the group consisting of primer binding sites, RNApromoters, or a combination of both.
 514. The process of claim 513,wherein said RNA promoters comprise phage promoters.
 515. The process ofclaim 514, wherein said phage promoters are selected from the groupconsisting of T3, T7 and SP6.
 516. The process of claim 497, whereinsaid hybridized nucleic acid copies further comprise one or moresignaling entities attached or incorporate thereto.
 517. The process ofclaim 516, wherein said signaling entities generate a signal directly orindirectly.
 518. The process of claim 517, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 519. The process of claim 517, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 520. The process ofclaim 519, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 521. The process of claim 497, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 522. The process of claim 497, wherein saidligating means comprise T4 DNA ligase.
 523. The process of claim 497,further comprising the step of separating the first copies obtained fromstep c) from their templates and repeating step b).
 524. The process ofclaim 497, further comprising the step of separating the extended secondset of primers obtained from step f) from their templates and repeatingstep e).
 525. The process of claim 497, wherein step g) is carried outrepeatedly.
 526. The process of claim 497, wherein said means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 527. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers and a second set of primers,wherein said second set comprises at least one production center; (iv) aset of oligonucleotides or polynucleotides complementary to at least onesegment or sequence of said second set of primers; and (v) means forligating said set of oligonucleotides or polynucleotides (iv); b)contacting said library of nucleic acid analytes with said first set ofprimers to form more than one first bound entity; c) extending saidbound first set of primers by means of template sequences provided bysaid nucleic acid analytes to form first copies of said analytes; d)ligating said set of oligonucleotides or polynucleotides a) (iv) to the3′ end of said first copies formed in step c) to form more than oneligated product; e) contacting said ligated product with said second setof primers to form more than one second bound entity; f) extending saidbound second set of primers by means of template sequences provided bysaid ligated products formed in step d) to form more than one complexcomprising said ligated products and said extended second set ofprimers; g) synthesizing from a production center in said second set ofprimers in said complexes one or more nucleic acid copies underisothermal or isostatic conditions; h) hybridizing said nucleic acidcopies formed in step g) to said array of nucleic acids provided in stepa) (i); and i) detecting or quantifying any of said hybridized copiesobtained in step h).
 528. The process of claim 527, wherein said nucleicacid array comprises members selected from the group consisting of DNA,RNA and analogs thereof.
 529. The process of claim 528, wherein saidanalogs comprise PNA.
 530. The process of claims 528 or 529, whereinsaid nucleic acids or analogs are modified on any one of the sugar,phosphate or base moieties.
 531. The process of claim 527, wherein saidnucleic acid array is fixed or immobilized to a solid support.
 532. Theprocess of claim 531, wherein said solid support is porous ornon-porous.
 533. The process of claim 532, wherein said porous solidsupport is selected from the group consisting of polyacrylamide andagarose.
 534. The process of claim 532, wherein said non-porous solidsupport comprises glass or plastic.
 535. The process of claim 531,wherein said solid support is transparent, translucent, opaque orreflective.
 536. The process of claim 531, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.537. The process of claim 536, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 538. The process of claim 527, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 539. The process ofclaim 527, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 540. Theprocess of claim 527, wherein said first set of primers comprise one ormore sequences which are complementary to inherent universal detectiontargets (UDTs).
 541. The process of claim 527, wherein said inherentUDTs are selected from the group consisting of 3′ poly A segments,consensus sequences, and a combination of both.
 542. The process ofclaim 541, wherein said consensus sequences is selected from the groupconsisting of signal sequences for poly A addition, splicing elements,multicopy repeats, and a combination of any of the foregoing.
 543. Theprocess of claim 527, wherein said production center is selected fromthe group consisting of primer binding sites, RNA promoters, or acombination of both.
 544. The process of claim 543, wherein said RNApromoters comprise phage promoters.
 545. The process of claim 544,wherein said phage promoters are selected from the group consisting ofT3, T7 and SP6.
 546. The process of claim 527, wherein said hybridizednucleic acid copies further comprise one or more signaling entitiesattached or incorporated thereto.
 547. The process of claim 546, whereinsaid signaling entities generate a signal directly or indirectly. 548.The process of claim 547, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 549. The process of claim 547, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 550. The process ofclaim 549, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 551. The process of claim 527, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 552. The process of claim 527, wherein saidligating means comprise T4 DNA ligase.
 553. The process of claim 527,wherein said means for synthesizing nucleic acid copies under isothermalor isostatic conditions is carried out by one or more members selectedfrom the group consisting of RNA transcription, strand displacementamplification and secondary structure amplification.
 554. The process ofclaim 527, further comprising the step of separating the first copiesobtained from step c) from their templates and repeating step b). 555.The process of claim 527, further comprising the step of separating theextended second set of primers obtained from step f) from theirtemplates and repeating step e).
 556. The process of claim 527, whereinstep g) is carried out repeatedly.
 557. The process of claim 527,wherein said means for synthesizing nucleic acid copies under isothermalor isostatic conditions is carried out by one or more members selectedfrom the group consisting of RNA transcription, strand displacementamplification and secondary structure amplification.
 558. A process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers, a second set of primers and athird set of primers wherein said third set comprises at least oneproduction center; and b) contacting said library of nucleic acidanalytes with said first set of primers to form a first set of boundprimers; c) extending said first set of bound primers by means oftemplate sequences provided by said nucleic acid analytes to form firstcopies of said analytes; d) contacting said extended first copies withsaid second set of primers to form a second set of bound primers; e)extending said second set of bound primers by means of templatesequences provided by said extended first copies to form second copiesof said nucleic acid analytes; f) contacting said second copies withsaid third set of primers to form more than one third bound entity toform a third set of bound primers; g) extending said third set of boundprimers by means of template sequences provided by said extended secondset of primers to form a hybrid comprising a second copy, a third copyand at least one production center; h) synthesizing from said productioncenter in said second set of primers in said complexes one or morenucleic acid copies under isothermal or isostatic conditions; i)hybridizing said nucleic acid copies formed in step i) to said array ofnucleic acids provided in step a) (i); and j) detecting or quantifyingany of said hybridized copies obtained in step i).
 559. The process ofclaim 558, wherein said nucleic acid array comprises members selectedfrom the group consisting of DNA, RNA and analogs thereof.
 560. Theprocess of claim 559, wherein said analogs comprise PNA.
 561. Theprocess of claims 559 or 560, wherein said nucleic acids or analogs aremodified on any one of the sugar, phosphate or base moieties.
 562. Theprocess of claim 558, wherein said nucleic acid array is fixed orimmobilized to a solid support.
 563. The process of claim 562, whereinsaid solid support is porous or non-porous.
 564. The process of claim563, wherein said porous solid support is selected from the groupconsisting of polyacrylamide and agarose.
 565. The process of claim 563,wherein said non-porous solid support comprises glass or plastic. 566.The process of claim 562, wherein said solid support is transparent,translucent, opaque or reflective.
 567. The process of claim 562,wherein said nucleic acids are directly or indirectly fixed orimmobilized to said solid support.
 568. The process of claim 562,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 569. Theprocess of claim 558, wherein said library of nucleic acid analytes isderived from a biological source selected from the group consisting oforgans, tissues and cells.
 570. The process of claim 558, wherein saidlibrary of nucleic acids analytes are derived from the group consistingof genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 571. The process of claim 558,wherein said first set of primers comprise one or more sequences whichare complementary to inherent universal detection targets (UDTs). 572.The process of claim 558, wherein said inherent UDTs are selected fromthe group consisting of 3′ poly A segments, consensus sequences, and acombination of both.
 573. The process of claim 572, wherein saidconsensus sequences is selected from the group consisting of signalsequences for poly A addition, splicing elements, multicopy repeats, anda combination of any of the foregoing.
 574. The process of claim 558,wherein said second set of primers are random primers.
 575. The processof claim 558, further comprising the step c′) of adding a primer bindingsite after step c).
 576. The process of claim 575, wherein said secondset of primers are complementary to said primer binding site.
 577. Theprocess of claim 575, wherein said primer binding site is added by meansof T4 DNA ligase or terminal transferase.
 578. The process of claim 558,wherein said production center is selected from the group consisting ofprimer binding sites, RNA promoters, or a combination of both.
 579. Theprocess of claim 578, wherein said RNA promoters comprise phagepromoters.
 580. The process of claim 579, wherein said phage promotersare selected from the group consisting of T3, T7 and SP6.
 581. Theprocess of claim 558, wherein said hybridized nucleic acid copiesfurther comprise one or more signaling entities attached or incorporatedthereto.
 582. The process of claim 581, wherein said signaling entitiesgenerate a signal directly or indirectly.
 583. The process of claim 582,wherein said direct signal generation is selected from the groupconsisting of a fluorescent compound, a phosphorescent compound, achemiluminescent compound, a chelating compound, an electron densecompound, a magnetic compound, an intercalating compound, an energytransfer compound and a combination of any of the foregoing.
 584. Theprocess of claim 582, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 585. The process of claim 584, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 586.The process of claim 558, wherein said polymerizing means are selectedfrom the group consisting of E. coli DNA Pol I, Klenow fragment of E.coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNApolymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.
 587. The process of claim 558, further comprising thestep of separating the first copies obtained from step c) from theirtemplates and repeating step b).
 588. The process of claim 558, furthercomprising the step of separating the extended second set of primersobtained from step f) from their templates and repeating step e). 589.The process of claim 558, wherein step g) is carried out repeatedly.590. The process of claim 558, further comprising the step f′) ofseparating said extended second set of primers obtained in step e). 591.The process of claim 558, further comprising the step of separating thefirst copies obtained from step c) from their templates and repeatingstep b).
 592. The process of claim 558, further comprising the step ofseparating the extended second set of primers obtained from step f) fromtheir templates and repeating step e).
 593. The process of claim 558,wherein step g) is carried out repeatedly.
 594. The process of claim558, wherein said means for synthesizing nucleic acid copies underisothermal or isostatic conditions is carried out by one or more membersselected from the group consisting of RNA transcription, stranddisplacement amplification and secondary structure amplification. 595.The process of claim 594, wherein said second set of primers comprise atleast one production center which differs in nucleotide sequence fromsaid production center in the third set of primers.
 596. A process fordetecting or quantifying more than one nucleic acid of interest in alibrary comprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers and a second set of primers,wherein said first set of primers are fixed or immobilized to a solidsupport, and wherein said second set of primers comprises at least twosegments, the first segment at the 3′ end comprising random sequences,and the second segment comprising at least one production center; (iv)means for synthesizing nucleic acid copies under isothermal or isostaticconditions; b) contacting said library of nucleic acid analytes withsaid first set of primers to form more than one first bound entity; c)extending said bound first set of primers by means of template sequencesprovided by said nucleic acid analytes to form first copies of saidanalytes; d) contacting said extended first copies with said second setof primers to form more than one second bound entity; e) extending saidbound second set of primers by means of template sequences provided bysaid extended first copies to form more than one complex comprisingextended first copies and extended second set of primers; f)synthesizing from a production center in said second set of primers insaid complexes one or more nucleic acid copies under isothermal orisostatic conditions; g) hybridizing said nucleic acid copies formed instep f) to said array of nucleic acids provided in step a) (i); and h)detecting or quantifying any of said hybridized copies obtained in stepg).
 597. The process of claim 596, wherein said solid support comprisesbeads.
 598. The process of claim 597, wherein said beads are magnetic.599. The process of claim 596, wherein said nucleic acid array comprisesmembers selected from the group consisting of DNA, RNA and analogsthereof.
 600. The process of claim 599, wherein said analogs comprisePNA.
 601. The process of claims 599 or 600, wherein said nucleic acidsor analogs are modified on any one of the sugar, phosphate or basemoieties.
 602. The process of claim 596, wherein said nucleic acid arrayis fixed or immobilized to a solid support.
 603. The process of claim602, wherein said solid support is porous or non-porous.
 604. Theprocess of claim 603, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 605. The process ofclaim 603, wherein said non-porous solid support comprises glass orplastic.
 606. The process of claim 602, wherein said solid support istransparent, translucent, opaque or reflective.
 607. The process ofclaim 602, wherein said nucleic acids are directly or indirectly fixedor immobilized to said solid support.
 608. The process of claim 607,wherein said nucleic acids are indirectly fixed or immobilized to saidsolid support by means of a chemical linker or linkage arm.
 609. Theprocess of claim 596, wherein said library of nucleic acid analytes isderived from a biological source selected from the group consisting oforgans, tissues and cells.
 610. The process of claim 596, wherein saidlibrary of nucleic acids analytes are derived from the group consistingof genomic DNA, episomal DNA, unspliced RNA, mRNA, rRNA, snRNA and acombination of any of the foregoing.
 611. The process of claim 596,wherein said first set of primers comprise one or more sequences whichare complementary to inherent universal detection targets (UDTs). 612.The process of claim 596, wherein said inherent UDTs are selected fromthe group consisting of 3′ poly A segments, consensus sequences, and acombination of both.
 613. The process of claim 612, wherein saidconsensus sequences is selected from the group consisting of signalsequences for poly A addition, splicing elements, multicopy repeats, anda combination of any of the foregoing.
 614. The process of claim 596,wherein said production center is selected from the group consisting ofprimer binding sites, RNA promoters, or a combination of both.
 615. Theprocess of claim 614, wherein said RNA promoters comprise phagepromoters.
 616. The process of claim 615, wherein said phage promotersare selected from the group consisting of T3, T7 and SP6.
 617. Theprocess of claim 596, wherein said hybridized nucleic acid copiesfurther i comprise one or more signaling entities attached orincorporated thereto.
 618. The process of claim 617, wherein saidsignaling entities generate a signal directly or indirectly.
 619. Theprocess of claim 618, wherein said direct signal generation is selectedfrom the group consisting of a fluorescent compound, a phosphorescentcompound, a chemiluminescent compound, a chelating compound, an electrondense compound, a magnetic compound, an intercalating compound, anenergy transfer compound and a combination of any of the foregoing. 620.The process of claim 618, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 621. The process of claim 620, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 622.The process of claim 596, wherein said polymerizing means are selectedfrom the group consisting of E. coli DNA Pol I, Klenow fragment of E.coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNApolymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.
 623. The process of claim 596, further comprising thestep of separating the first copies obtained from step c) from theirtemplates and repeating step b).
 624. The process of claim 596, furthercomprising the step of separating the extended second set of primersobtained from step f) from their templates and repeating step e). 625.The process of claim 596, wherein step g) is carried out repeatedly.626. The process of claim 596, wherein said means for synthesizingnucleic acid copies under isothermal or isostatic conditions is carriedout by one or more members selected from the group consisting of RNAtranscription, strand displacement amplification and secondary structureamplification.
 627. A process for detecting or quantifying more than onenucleic acid of interest in a library comprising the steps of: a)providing: (i) an array of fixed or immobilized nucleic acids identicalor complementary in part or whole to sequences of said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; (iii)polymerizing means for synthesizing nucleic acid copies of said nucleicacid analytes, said polymerizing means comprising a first set of primersand a second set of primers, wherein said first set of primers are fixedor immobilized to a solid support, and wherein said first set of primerscomprise at least one production center; and (iv) means for synthesizingnucleic acid copies under isothermal or isostatic conditions; b)contacting said library of nucleic acid analytes with said first set ofprimers to form more than one first bound entity; c) extending saidbound first set of primers by means of template sequences provided bysaid nucleic acid analytes to form first copies of said analytes; d)extending said first copies by means of at least four (4) or morenon-inherent homopolymeric nucleotides; e) contacting said extendedfirst copies with said second set of primers to form more than onesecond bound entity; f) extending said bound second set of primers bymeans of template sequences provided by said extended first copies toform more than one complex comprising extended first copies and extendedsecond set of primers; g) synthesizing from a production center in saidsecond set of primers in said complexes one or more nucleic acid copiesunder isothermal or isostatic conditions; h) hybridizing said nucleicacid copies formed in step g) to said array of nucleic acids provided instep a) (i); and i) detecting or quantifying any of said hybridizedcopies obtained in step h).
 628. The process of claim 627, wherein saidsolid support comprises beads.
 629. The process of claim 628, whereinsaid beads are magnetic.
 630. The process of claim 627, wherein saidnucleic acid array comprises members selected from the group consistingof DNA, RNA and analogs thereof.
 631. The process of claim 630, whereinsaid analogs comprise PNA.
 632. The process of claims 630 or 631,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 633. The process of claim 627,wherein said nucleic acid array is fixed or immobilized to a solidsupport.
 634. The process of claim 633, wherein said solid support isporous or non-porous.
 635. The process of claim 634, wherein said poroussolid support is selected from the group consisting of polyacrylamideand agarose.
 636. The process of claim 634, wherein said non-poroussolid support comprises glass or plastic.
 637. The process of claim 633,wherein said solid support is transparent, translucent, opaque orreflective.
 638. The process of claim 633, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.639. The process of claim 638, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 640. The process of claim 627, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 641. The process ofclaim 627, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 642. Theprocess of claim 627, wherein said first set of primers further compriseone or more sequences complementary to inherent universal detectiontargets (UDTs).
 643. The process of claim 627, wherein said inherent UDTis selected from the group consisting of 3′ poly A segments, consensussequences, and a combination of both.
 644. The process of claim 643,wherein said consensus sequences is selected from the group consistingof signal sequences for poly A addition, splicing elements, multicopyrepeats, and a combination of any of the foregoing.
 645. The process ofclaim 627, wherein said production center is selected from the groupconsisting of primer binding sites, RNA promoters, or a combination ofboth.
 646. The process of claim 645, wherein said RNA promoters comprisephage promoters.
 647. The process of claim 646, wherein said phagepromoters are selected from the group consisting of T3, T7 and SP6. 648.The process of claim 627, wherein said extending step d), the four ormore non-inherent homopolymeric nucleotides are added by terminaltransferase.
 649. The process of claim 627, wherein said hybridizednucleic acid copies further comprise one or more signaling entitiesattached or incorporated thereto.
 650. The process of claim 649, whereinsaid signaling entities generate a signal directly or indirectly. 651.The process of claim 650, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 652. The process of claim 650, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 653. The process ofclaim 652, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 654. The process of claim 627, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 655. The process of claim 627, furthercomprising the step of separating the first copies obtained from step c)from their templates and repeating step b).
 656. The process of claim627, further comprising the step of separating the extended second setof primers obtained from step f) from their templates and repeating stepe).
 657. The process of claim 627, wherein step g) is carried outrepeatedly.
 658. The process of claim 627, wherein said means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 659. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers and a second set of primers,wherein said first set of primers are fixed or immobilized to a solidsupport, and wherein said first set comprises at least one productioncenter; (iv) a set of oligonucleotides or polynucleotides complementaryto at least one segment or sequence of said second set of primers; and(v) means for ligating said set of oligonucleotides or polynucleotides(iv); b) contacting said library of nucleic acid analytes with saidfirst set of primers to form more than one first bound entity; c)extending said bound first set of primers by means of template sequencesprovided by said nucleic acid analytes to form first copies of saidanalytes; d) ligating said set of oligonucleotides or polynucleotides a)(iv) to the 3′ end of said first copies formed in step c) to form morethan one ligated product; e) contacting said ligated product with saidsecond set of primers to form more than one second bound entity; f)extending said bound second set of primers by means of templatesequences provided by said ligated products formed in step d) to formmore than one complex comprising said ligated products and said extendedsecond set of primers; g) synthesizing from a production center in saidsecond set of primers in said complexes one or more nucleic acid copiesunder isothermal or isostatic conditions; h) hybridizing said nucleicacid copies formed in step g) to said array of nucleic acids provided instep a) (i); and i) detecting or quantifying any of said hybridizedcopies obtained in step h).
 660. The process of claim 659, wherein saidsolid support comprises beads.
 661. The process of claim 660, whereinsaid beads are magnetic.
 662. The process of claim 659, wherein saidnucleic acid array comprises members selected from the group consistingof DNA, RNA and analogs thereof.
 663. The process of claim 662, whereinsaid analogs comprise PNA.
 664. The process of claims 662 or 663,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 665. The process of claim 659,wherein said nucleic acid array is fixed or immobilized to a solidsupport.
 666. The process of claim 665, wherein said solid support isporous or non-porous.
 667. The process of claim 666, wherein said poroussolid support is selected from the group consisting of polyacrylamideand agarose.
 668. The process of claim 666, wherein said non-poroussolid support comprises glass or plastic.
 669. The process of claim 665,wherein said solid support is transparent, translucent, opaque orreflective.
 670. The process of claim 665, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.671. The process of claim 665, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 672. The process of claim 659, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 673. The process ofclaim 659, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 674. Theprocess of claim 659, wherein said first set of primers comprise one ormore sequences which are complementary to inherent universal detectiontargets (UDTs).
 675. The process of claim 659, wherein said inherentUDTs are selected from the group consisting of 3′ poly A segments,consensus sequences, and a combination of both.
 676. The process ofclaim 675, wherein said consensus sequences is selected from the groupconsisting of signal sequences for poly A addition, splicing elements,multicopy repeats, and a combination of any of the foregoing.
 677. Theprocess of claim 659, wherein said production center is selected fromthe group consisting of primer binding sites, RNA promoters, or acombination of both.
 678. The process of claim 677, wherein said RNApromoters comprise phage promoters.
 679. The process of claim 678,wherein said phage promoters are selected from the group consisting ofT3, T7 and SP6.
 680. The process of claim 659, wherein said hybridizednucleic acid copies further comprise one or more signaling entitiesattached or incorporated thereto.
 681. The process of claim 680, whereinsaid signaling entities generate a signal directly or indirectly. 682.The process of claim 681, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 683. The process of claim 682, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 684. The process ofclaim 683, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 685. The process of claim 659, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 686. The process of claim 659, wherein saidligating means comprise T4 DNA ligase.
 687. The process of claim 659,further comprising the step of separating the first copies obtained fromstep c) from their templates and repeating step b).
 688. The process ofclaim 659, further comprising the step of separating the extended secondset of primers obtained from step f) from their templates and repeatingstep e).
 689. The process of claim 659, wherein step g) is carried outrepeatedly.
 690. The process of claim 659, wherein said means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 691. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers and a second set of primers,wherein said first set of primers are fixed or immobilized to a solidsupport, and wherein said second set comprises at least one productioncenter; (iv) a set of oligonucleotides or polynucleotides complementaryto at least one segment or sequence of said second set of primers; and(v) means for ligating said set of oligonucleotides or polynucleotides(iv); b) contacting said library of nucleic acid analytes with saidfirst set of primers to form more than one first bound entity; c)extending said bound first set of primers by means of template sequencesprovided by said nucleic acid analytes to form first copies of saidanalytes; d) ligating said set of oligonucleotides or polynucleotides a)(iv) to the 3′ end of said first copies formed in step c) to form morethan one ligated product; e) contacting said ligated product with saidsecond set of primers to form more than one second bound entity; f)extending said bound second set of primers by means of templatesequences provided by said ligated products formed in step d) to formmore than one complex comprising said ligated products and said extendedsecond set of primers; g) synthesizing from a production center in saidsecond set of primers in said complexes one or more nucleic acid copiesunder isothermal or isostatic conditions; h) hybridizing said nucleicacid copies formed in step g) to said array of nucleic acids provided instep a) (i); and i) detecting or quantifying any of said hybridizedcopies obtained in step h).
 692. The process of claim 691, furthercomprising the step of separating the first copies obtained from step c)from their templates and repeating step b).
 693. The process of claim691, further comprising the step of separating the extended second setof primers obtained from step f) from their templates and repeating stepe).
 694. The process of claim 691, wherein step g) is carried outrepeatedly.
 695. The process of claim 691, wherein said means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 696. A process for detecting orquantifying more than one nucleic acid of interest in a librarycomprising the steps of: a) providing: (i) an array of fixed orimmobilized nucleic acids identical or complementary in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; and (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes, said polymerizingmeans comprising a first set of primers, a second set of primers and athird set of primers, wherein said first set of primers are fixed orimmobilized to a solid support, and wherein said third set comprises atleast one production center; and b) contacting said library of nucleicacid analytes with said first set of primers to form more than one firstbound entity; c) extending said bound first set of primers by means oftemplate sequences provided by said nucleic acid analytes to form firstcopies of said analytes; d) contacting said extended first copies withsaid second set of primers to form more than one second bound entity; e)extending said bound second set of primers by means of templatesequences provided by said extended first copies to form an extendedsecond set of primers; f) separating said extended second set of primersobtained in step e) g) contacting said extended second set of primerswith said third set of primers to form more than one third bound entity;h) extending said third bound entity by means of template sequencesprovided by said extended second set of primers to form more than onecomplex comprising said extended third bound entity and said extendedset of primers; i) synthesizing from a production center in said secondset of primers in said complexes one or more nucleic acid copies underisothermal or isostatic conditions; j) hybridizing said nucleic acidcopies formed in step i) to said array of nucleic acids provided in stepa) (i); and k) detecting or quantifying any of said hybridized copiesobtained in step j)
 697. The process of claim 696, wherein said solidsupport comprises beads.
 698. The process of claim 697, wherein saidbeads are magnetic.
 699. The process of claim 696, wherein said nucleicacid array comprises members selected from the group consisting of DNA,RNA and analogs thereof.
 700. The process of claim 699, wherein saidanalogs comprise PNA.
 701. The process of claims 699 or 700, whereinsaid nucleic acids or analogs are modified on any one of the sugar,phosphate or base moieties.
 702. The process of claim 696, wherein saidnucleic acid array is fixed or immobilized to a solid support.
 703. Theprocess of claim 702, wherein said solid support is porous ornon-porous.
 704. The process of claim 703, wherein said porous solidsupport is selected from the group consisting of polyacrylamide andagarose.
 705. The process of claim 703, wherein said non-porous solidsupport comprises glass or plastic.
 706. The process of claim 703,wherein said solid support is transparent, translucent, opaque orreflective.
 707. The process of claim 703, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.708. The process of claim 707, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 709. The process of claim 696, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 710. The process ofclaim 696, wherein said library of nucleic acids analytes are derivedfrom the group consisting of genomic DNA, episomal DNA, unspliced RNA,mRNA, rRNA, snRNA and a combination of any of the foregoing.
 711. Theprocess of claim 696, wherein said first set of primers comprise one ormore sequences which are complementary to inherent universal detectiontargets (UDTs).
 712. The process of claim 696, wherein said inherentUDTs are selected from the group consisting of 3′ poly A segments,consensus sequences, and a combination of both.
 713. The process ofclaim 712, wherein said consensus sequences is selected from the groupconsisting of signal sequences for poly A addition, splicing elements,multicopy repeats, and a combination of any of the foregoing.
 714. Theprocess of claim 696, wherein said second set of primers are randomprimers.
 715. The process of claim 696, further comprising the step c′)of adding a primer binding site after step c).
 716. The process of claim715, wherein said second set of primers are complementary to said primerbinding site.
 717. The process of claim 715, wherein said primer bindingsite is added by means of T4 DNA ligase or terminal transferase. 718.The process of claim 696, wherein said production center is selectedfrom the group consisting of primer binding sites, RNA promoters, or acombination of both.
 719. The process of claim 718, wherein said RNApromoters comprise phage promoters.
 720. The process of claim 719,wherein said phage promoters are selected from the group consisting ofT3, T7 and SP6.
 721. The process of claim 696, wherein said hybridizednucleic acid copies further comprise one or more signaling entitiesattached or incorporated thereto.
 722. The process of claim 721, whereinsaid signaling entities generate a signal directly or indirectly. 723.The process of claim 722, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 724. The process of claim 722, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 725. The process ofclaim 724, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 726. The process of claim 696, wherein saidpolymerizing means are selected from the group consisting of E. coli DNAPol I, Kienow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 727. The process of claim 696, furthercomprising the step of separating the first copies obtained from step c)from their templates and repeating step b).
 728. The process of claim696, further comprising the step of separating the extended second setof primers obtained from step f) from their templates and repeating stepe).
 729. The process of claim 696, wherein step g) is carried outrepeatedly.
 730. The process of claim 696, wherein said means forsynthesizing nucleic acid copies under isothermal or isostaticconditions is carried out by one or more members selected from the groupconsisting of RNA transcription, strand displacement amplification andsecondary structure amplification.
 731. The process of claim 696,wherein said second set of primers comprise at least one productioncenter which differs in nucleotide sequence from said production centerin the third set of primers.
 732. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing (i) an array of fixed or immobilized nucleic acidsidentical in part or whole to sequences of said nucleic acids ofinterest; (ii) a library of nucleic acid analytes which may contain thenucleic acids of interest sought to be detected or quantified; and (iii)polymerizing means for synthesizing nucleic acid copies of said nucleicacid analytes said polymerizing means comprising a first set of primers;b) contacting said nucleic acid analytes with said first set of primersto form a first bound entity; c) extending said bound set of first setof primers by means of template sequences provided by said nucleic acidanalytes to form first nucleic acid copies of said analytes; d)separating said first nucleic acid copies from the said analytes; e)repeating steps b), c) and d) until a desirable amount of first nucleicacid copies have been synthesized; f) hybridizing said nucleic nucleicacid copies formed in step e) to said array of nucleic acids provided instep (i); and g) detecting or quantifying any of said hybridized firstnucleic acid copies obtained in step f).
 733. The process of claim 732,wherein said nucleic acid array is selected from the group consisting ofDNA, RNA and analogs thereof.
 734. The process of claim 4, wherein saidanalogs comprise PNA.
 735. The process of claims 733 or 734, whereinsaid nucleic acids or analogs are modified on any one of the sugar,phosphate or base moieties.
 736. The process of claim 732, wherein saidarray of nucleic acids are fixed or immobilized to a solid support. 737.The process of claim 736, wherein said solid support is porous ornon-porous.
 738. The process of claim 737, wherein said porous solidsupport is selected from the group consisting of polyacrylamide andagarose.
 739. The process of claim 736, wherein said non-porous solidsupport comprises glass or plastic.
 740. The process of claim 736,wherein said solid support is transparent, translucent, opaque orreflective.
 741. The process of claim 736, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.742. The process of claim 741, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 743. The process of claim 732, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 744. The process ofclaim 732, wherein said nucleic acid analytes are selected from thegroup consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.
 745. The processof claim 732, wherein said nucleic acid analytes comprise an inherentUDT selected from the group consisting of poly T segments, secondarystructures, consensus sequences, and a combination of any of theforegoing.
 746. The process of claim 745, wherein said consensussequences is selected from the group consisting of signal sequences forpoly A addition, splicing elements, multicopy repeats, and a combinationof any of the foregoing.
 747. The process of claim 732, furthercomprising the step of adding one or more non-inhererent UDTs to saidnucleic acid analytes or said first copies by an enzymatic meansselected from the group consisting of poly A polymerase, terminaltransferase, T4 DNA ligase, T4 RNA ligase and a combination of any ofthe foregoing.
 748. The process of claim 732, wherein said providing orcontacting steps, the first set of primers comprise one or more UDTs.749. The process of claim 732, wherein said polymerizing means comprisesan enzyme selected from the group consisting of E. coli DNA Pol I,Klenow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 750. The process of claim 749, wherein anadditional amount of enzyme is added after step d) or after repeatingstep d).
 751. The process of claim 732, wherein said hybridized nucleicacid copies further comprise one or more signaling entities attached orincorporated thereto.
 752. The process of claim 748, wherein said UDEgenerates a signal directly or indirectly.
 753. The process of claim752, wherein said direct signal generation is selected from the groupconsisting of a fluorescent compound, a phosphorescent compound, achemiluminescent compound, a chelating compound, an electron densecompound, a magnetic compound, an intercalating compound, an energytransfer compound and a combination of any of the foregoing.
 754. Theprocess of claim 752, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 755. The process of claim 754, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction.
 756. Aprocess for detecting or quantifying more than one nucleic acid ofinterest in a library comprising the steps of: a) providing (i) an arrayof fixed or immobilized nucleic acids identical in part or whole tosequences of said nucleic acids of interest; (ii) a library of nucleicacid analytes which may contain the nucleic acids of interest sought tobe detected or quantified; (iii) polymerizing means for synthesizingnucleic acid copies of said nucleic acid analytes said polymerizingmeans comprising a first set of primers and a second set of primers;(iv) means for addition of sequences to the 3′ end of nucleic acids; b)contacting said nucleic acid analytes with said first set of primer toform a first bound entity; c) extending said bound set of first set ofprimers by means of template sequences provided by said nucleic acidanalytes to form first nucleic acid copies of said analytes; d)extending said first nucleic copies by the addition of non-templatederived sequences to the 3′ end of said first nucleic acid copies e)contacting said extended first nucleic acid copies with said second setof primers to form a second bound entity; f) extending said bound set ofsecond set of primers by means of template sequences provided by saidextended first nucleic acid copies to form second nucleic acid copies;g) separating said second nucleic acid copies from the extended firstnucleic acid copies; h) repeating steps e), f) and g) until a desirableamount of second nucleic acid copies have been synthesized; i)hybridizing said second nucleic acid copies formed in step h) to saidarray of nucleic acids provided in step (i); and j) detecting orquantifying any of said hybridized second nucleic acid copies obtainedin step i).
 757. The process of claim 756, wherein said nucleic acidarray is selected from the group consisting of DNA, RNA and analogsthereof.
 758. The process of claim 757, wherein said analogs comprisePNA.
 759. The process of claims 757 or 758, wherein said nucleic acidsor analogs are modified on any one of the sugar, phosphate or basemoieties.
 760. The process of claim 756, wherein said solid support isporous or non-porous.
 761. The process of claim 760, wherein said poroussolid support is selected from the group consisting of polyacrylamideand agarose.
 762. The process of claim 760, wherein said non-poroussolid support comprises glass or plastic.
 763. The process of claim 758,wherein said solid support is transparent, translucent, opaque orreflective.
 764. The process of claim 756, wherein said nucleic acidsare directly or indirectly fixed or immobilized to said solid support.765. The process of claim 764, wherein said nucleic acids are indirectlyfixed or immobilized to said solid support by means of a chemical linkeror linkage arm.
 766. The process of claim 758, wherein said library ofnucleic acid analytes is derived from a biological source selected fromthe group consisting of organs, tissues and cells.
 767. The process ofclaim 758, wherein said nucleic acid analytes are selected from thegroup consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.
 768. The processof claim 758, wherein said nucleic acid analytes comprise an inherentUDT selected from the group consisting of poly T segments, secondarystructures, consensus sequences, and a combination of any of theforegoing.
 769. The process of claim 768, wherein said consensussequences is selected from the group consisting of signal sequences forpoly A addition, splicing elements, multicopy repeats, and a combinationof any of the foregoing.
 770. The process of claim 758, furthercomprising the step of adding one or more non-inhererent UDTs to saidnucleic acid analytes, said first copies or said second copies by anenzymatic means selected from the group consisting of poly A polymerase,terminal transferase, T4 DNA ligase, T4 RNA ligase and a combination ofany of the foregoing.
 771. The process of claim 758, wherein saidproviding or contacting steps, the first set of primers or the secondset of primers or both comprise one or more UDTs.
 772. The process ofclaim 758, wherein said extending step d) is carried out by an enzymaticmeans selected from the group consisting of terminal transferase, T4 DNAligase, T4 RNA ligase, and a combination of any of the foregoing. 773.The process of claim 758, wherein said polymerizing means comprises anenzyme selected from the group consisting of E. coli DNA Pol I, Klenowfragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNA polymerase,Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase, ALV reversetranscriptase, MuLV reverse transcriptase, RSV reverse transcriptase,HIV-1 reverse transcriptase, HIV-2 reverse transcriptase, Sensiscriptand Omniscript.
 774. The process of claim 773, wherein following one ormore separation steps an additional amount of enzyme is added.
 775. Theprocess of claim 758, wherein said hybridized nucleic acid copiesfurther comprise one or more signaling entities attached or incorporatedthereto.
 776. The process of claim 775, wherein said signaling entitiesgenerate a signal directly or indirectly.
 777. The process of claim 776,wherein said direct signal generation is selected from the groupconsisting of a fluorescent compound, a phosphorescent compound, achemiluminescent compound, a chelating compound, an electron densecompound, a magnetic compound, an intercalating compound, an energytransfer compound and a combination of any of the foregoing.
 778. Theprocess of claim 776, wherein said indirect signal generation isselected from the group consisting of an antibody, an antigen, a hapten,a receptor, a hormone, a ligand, an enzyme and a combination of any ofthe foregoing.
 779. The process of claim 778, wherein said enzymecatalyzes a reaction selected from the group consisting of a fluorogenicreaction, a chromogenic reaction and a chemiluminescent reaction. 780.The process of claim 756, which comprises the additional steps of k)separating the first nucleic copies produced in step c) of claim 1750from said analytes l) repeating steps b) c) and k) until a desirableamount of first nucleic acid copies have been synthesized.
 781. Theprocess of claim 756, which comprises the additional steps of 1)separating the extended first nucleic copies produced in step d) ofclaim 1750 from said analytes and m) repeating steps b), c), d) and 1)until a desirable amount of extended first nucleic acid copies have beensynthesized.
 782. The process of claim 756, wherein said first set ofprimers are attached to a solid support.
 783. The process of claim 782,wherein said solid support comprises beads.
 784. The process of claim783, wherein said beads are magnetic.
 785. A composition of matter thatcomprises an array of solid surfaces comprising discrete areas; whereinat least two of said discrete areas each comprises: a first set ofnucleic acid primers; and a second set of nucleic acid primers; whereinthe nucleotide sequences in said first set of nucleic acid primers aredifferent from the nucleotide sequences in said second set of nucleicacid primers; wherein the nucleotide sequences of a first set of nucleicacid primers of a first discrete area and the nucleotide sequences of afirst set of nucleic acid primers of a second discrete area differ fromeach other by at least one base; and wherein the nucleotide sequences ofthe second set of nucleic acid primers of a first discrete area and thenucleotide sequences of the second set of nucleic acid primers of asecond discrete area are substantially the same or identical.
 786. Thecomposition of claim 785, wherein said array of solid surfaces has beendesigned/synthesized such that D1 is less than D2, said D1 being thephysical distance on said array between a nucleic acid primer that ispart of a first set of an area and the nucleic acid primer is part of asecond set of the same area, and D2 being the physical distance in anucleic acid in a sample between the sequence of a primer binding sitein said nucleic acid in a sample for the nucleic acid primer of thefirst set and the complement of the primer binding site in the saidnucleic acid in the sample for the nucleic acid primer in the secondset.
 787. The composition of claim 785, wherein said nucleic acidprimers are selected from the group consisting of DNA, RNA and analogsthereof.
 788. The composition of claim 787, wherein said analogscomprise PNA.
 789. The composition of claims 787 or 788, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 790. The composition of claim 785, wherein said solidsurfaces are porous or non-porous.
 791. The composition of claim 789,wherein said porous solid surfaces are selected from the groupconsisting of polyacrylamide and agarose.
 792. The composition of claim790, wherein said non-porous solid surfaces comprise glass or plastic.793. The composition of claim 785, wherein said solid surfaces aretransparent, translucent, opaque or reflective.
 794. The composition ofclaim 785, wherein nucleic acid primers are directly or indirectly fixedor immobilized to said solid surfaces.
 795. The composition of claim794, wherein said nucleic acid primers are indirectly fixed orimmobilized to said solid surfaces by means of a chemical linker orlinkage arm.
 796. A composition of matter that comprises an array ofsolid surfaces comprising a plurality of discrete areas; wherein atleast two of said discrete areas each comprises: a first set of nucleicacid primers; and a second set of nucleic acid primers; wherein thenucleotide sequences in said first set of nucleic acid primers aredifferent from the nucleotide sequences in said second set of nucleicacid primers; wherein the nucleotide sequences of a first set of nucleicacid primers of a first discrete area and the nucleotide sequences of afirst set of nucleic acid primers of a second discrete area differsubstantially from each other; and wherein the nucleotide sequences ofthe second set of nucleic acid primers of a first discrete area and thenucleotide sequences of the second set of nucleic acid primers of asecond discrete area are substantially the same or identical.
 797. Thecomposition of claim 795, wherein said array of solid surfaces has beendesigned/synthesized such that D1 is less than D2, said D1 being thephysical distance on said array between a nucleic acid primer that ispart of a first set of an area and the nucleic acid primer is part of asecond set of the same area, and D2 being the physical distance in anucleic acid in a sample between the sequence of a primer binding sitein said nucleic acid in a sample for the nucleic acid primer of thefirst set and the complement of the primer binding site in the saidnucleic acid in the sample for the nucleic acid primer in the secondset.
 798. The composition of claim 796, wherein said nucleic acidprimers are selected from the group consisting of DNA, RNA and analogsthereof.
 799. The composition of claim 798, wherein said analogscomprise PNA.
 800. The composition of claims 798 or 799, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 801. The composition of claim 796, wherein said solidsurfaces are porous or non-porous.
 802. The composition of claim 801,wherein said porous solid surfaces are selected from the groupconsisting of polyacrylamide and agarose.
 803. The composition of claim796, wherein said non-porous solid surfaces comprise glass or plastic.804. The composition of claim 796, wherein said solid surfaces aretransparent, translucent, opaque or reflective.
 805. The composition ofclaim 796, wherein nucleic acid primers are directly or indirectly fixedor immobilized to said solid surfaces.
 806. The composition of claim805, wherein said nucleic acid primers are indirectly fixed orimmobilized to said solid surfacees by means of a chemical linker orlinkage arm.
 807. A process for producing two or more copies of nucleicacids of interest in a library comprising the steps of: a) providing:(i) an array of solid surfaces comprising a plurality of discrete areas;wherein at least two of said discrete areas each comprises: (1) a firstset of nucleic acid primers; and (2) a second set of nucleic acidprimers; wherein the nucleotide sequences in said first set of nucleicacid primers are different from the nucleotide sequences in said secondset of nucleic acid primers; wherein the nucleotide sequences of a firstset of nucleic acid primers of a first discrete area and the nucleotidesequences of a first set of nucleic acid primers of a second discretearea differ from each other by at least one base; and wherein thenucleotide sequences of the second set of nucleic acid primers of afirst discrete area and the nucleotide sequences of the second set ofnucleic acid primers of a second discrete area are substantially thesame or identical; (ii) a library of nucleic acid analytes which maycontain the nucleic acids of interest; (iii) polymerizing means forsynthesizing nucleic acid copies of said nucleic acids of interest; b)contacting a primer of said first set with a complementary sequence insaid nucleic acid of interest; c) extending said primer in the first setusing said nucleic acid of interest as a template to generate anextended first primer; d) contacting a primer in said second set with acomplementary sequence in said extended first primer; e) extending saidprimer in the second set using said extended first primer as a templateto generate an extended second primer; f) contacting a primer in thefirst set with a complementary sequence in said extended second primer;g) extending said primer in the first set using said extended secondprimer as a template to generate an extended first primer; and h)repeating steps d) through g) above one or more times.
 808. The processof claim 807, wherein said nucleic acid primers are selected from thegroup consisting of DNA, RNA and analogs thereof.
 809. The process ofclaim 808, wherein said analogs comprise PNA.
 810. The process of claims808 or 809, wherein said nucleic acids or analogs are modified on anyone of the sugar, phosphate or base moieties.
 811. The process of claim807, wherein said solid support is porous or non-porous.
 812. Theprocess of claim 811, wherein said porous solid support is selected fromthe group consisting of polyacrylamide and agarose.
 813. The process ofclaim 811, wherein said non-porous solid support comprises glass orplastic.
 814. The process of claim 807, wherein said solid support istransparent, translucent, opaque or reflective.
 815. The process ofclaim 807, wherein nucleic acid primers are directly or indirectly fixedor immobilized to said solid support.
 816. The process of claim 815,wherein said nucleic acid primers are indirectly fixed or immobilized tosaid solid support by means of a chemical linker or linkage arm. 817.The process of claim 807, wherein said library of analytes is derivedfrom a biological source selected from the group consisting of organs,tissues and cells.
 818. The process of claim 807, wherein said analytesare selected from the group consisting of genomic DNA, episomal DNA,unspliced RNA, mRNA, rRNA, snRNA and a combination of any of theforegoing.
 819. The process of claim 600, wherein said polymerizingmeans are selected from the group consisting of E. coli DNA Pol I,Kienow fragment of E. coli DNA Pol I, Bst DNA polymerase, Bca DNApolymerase, Taq DNA polymerase, Tth DNA Polymerase, T4 DNA polymerase,ALV reverse transcriptase, MuLV reverse transcriptase, RSV reversetranscriptase, HIV-1 reverse transcriptase, HIV-2 reverse transcriptase,Sensiscript and Omniscript.
 820. A process for detecting or quantifyingmore than one nucleic acid of interest in a library comprising the stepsof: a) providing: (i) an array of solid surfaces comprising a pluralityof discrete areas; wherein at least two of said discrete areas eachcomprises: (1) a first set of nucleic acid primers; and (2) a second setof nucleic acid primers; wherein the nucleotide sequences in said firstset of nucleic acid primers are different from the nucleotide sequencesin said second set of nucleic acid primers; wherein the nucleotidesequences of a first set of nucleic acid primers of a first discretearea and the nucleotide sequences of a first set of nucleic acid primersof a second discrete area differ from each other by at least one base;and wherein the nucleotide sequences of the second set of nucleic acidprimers of a first discrete area and the nucleotide sequences of thesecond set of nucleic acid primers of a second discrete area aresubstantially the same or identical; (ii) a library of nucleic acidanalytes which may contain the nucleic acids of interest; (iii)polymerizing means for synthesizing nucleic acid copies of said nucleicacids of interest; and (iv) non-radioactive signal generating meanscapable of being attached to or incorporated into nucleic acids; b)contacting a primer of said first set with a complementary sequence insaid nucleic acid of interest; c) extending said primer in the first setusing said nucleic acid of interest as a template to generate anextended first primer; d) contacting a primer in said second set with acomplementary sequence in said extended first primer; e) extending saidprimer in the second set using said extended first primer as a templateto generate an extended second primer; f) contacting a primer in thefirst set with a complementary sequence in said extended second primer;g) extending said primer in the first set using said extended secondprimer as a template to generate an extended first primer; h) repeatingsteps d) through g) above one or more times; and i) detecting orquantifying by means of said non-radioactive signal generating meansattached to or incorporated into any of said extended primers in stepsc), e), g), and h).
 821. The process of claim 820, wherein said nucleicacid primers are selected from the group consisting of DNA, RNA andanalogs thereof.
 822. The process of claim 821, wherein said analogscomprise PNA.
 823. The process of claims 821 or 822, wherein saidnucleic acids or analogs are modified on any one of the sugar, phosphateor base moieties.
 824. The process of claim 820, wherein said solidsupport is porous or non-porous.
 825. The process of claim 824, whereinsaid porous solid support is selected from the group consisting ofpolyacrylamide and agarose.
 826. The process of claim 824, wherein saidnon-porous solid support comprises glass or plastic.
 827. The process ofclaim 820, wherein said solid support is transparent, translucent,opaque or reflective.
 828. The process of claim 820, wherein nucleicacid primers are directly or indirectly fixed or immobilized to saidsolid support.
 829. The process of claim 828, wherein said nucleic acidprimers are indirectly fixed or immobilized to said solid support bymeans of a chemical linker or linkage arm.
 830. The process of claim820, wherein said library of analytes is derived from a biologicalsource selected from the group consisting of organs, tissues and cells.831. The process of claim 820, wherein said analytes are selected fromthe group consisting of genomic DNA, episomal DNA, unspliced RNA, mRNA,rRNA, snRNA and a combination of any of the foregoing.
 832. The processof claim 820, wherein said polymerizing means are selected from thegroup consisting of E. coli DNA Pol I, Klenow fragment of E. coli DNAPol I, Bst DNA polymerase, Bca DNA polymerase, Taq DNA polymerase, TthDNA Polymerase, T4 DNA polymerase, ALV reverse transcriptase, MuLVreverse transcriptase, RSV reverse transcriptase, HIV-1 reversetranscriptase, HIV-2 reverse transcriptase, Sensiscript and Omniscript.833. The process of claim 820, wherein said non-radioactive signalgenerating means are selected from the group consisting of labelednucleotides, intercalating dyes, universal detection elements and acombination of any of the foregoing.
 834. The process of claim 820,wherein said extended primers further comprise one or more signalingentities attached or incorporated thereto.
 835. The process of claim834, wherein said signaling entities generate a signal directly orindirectly.
 836. The process of claim 835, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 837. The process of claim 835, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 838. The process ofclaim 837, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 839. A composition of matter that comprisesan array of solid surfaces comprising a plurality of discrete areas;wherein at least two of said discrete areas comprise: a chimericcomposition comprising: a nucleic acid portion; and a non-nucleic acidportion; wherein said nucleic acid portion of a first discrete area hasthe same sequence as the nucleic acid portion of a second discrete area;and wherein said non-nucleic acid portion has a binding affinity foranalytes of interest.
 840. The composition of claim 839, wherein saidnucleic acid portion is selected from the group consisting of DNA, RNAand analogs thereof.
 841. The composition of claim 840, wherein saidanalogs comprise PNA.
 842. The composition of claims 840 or 841, whereinsaid nucleic acids or analogs are modified on any one of the sugar,phosphate or base moieties.
 843. The composition of claim 839, whereinsaid solid surfaces are porous or non-porous.
 844. The composition ofclaim 843, wherein said porous solid surfaces are selected from thegroup consisting of polyacrylamide and agarose.
 845. The composition ofclaim 843, wherein said non-porous solid surfaces comprise glass orplastic.
 846. The composition of claim 839, wherein said solid surfacesare transparent, translucent, opaque or reflective.
 847. The compositionof claim 839, wherein said nucleic acid portions are directly orindirectly fixed or immobilized to said solid surfaces.
 848. Thecomposition of claim 839, wherein said non-nucleic acid portions areselected from the group consisting of peptides, proteins, ligands,enzyme substrates, hormones, receptors, drugs and a combination of anyof the foregoing.
 849. A composition of matter that comprises an arrayof solid surfaces comprising a plurality of discrete areas; wherein atleast two of said discrete areas comprise: a chimeric compositionhybridized to complementary sequences of nucleic acids fixed orimmobilized to said discrete areas, wherein said chimeric compositioncomprises: a nucleic acid portion; and a non-nucleic acid portion; saidnucleic acid portion comprising at least one sequence, wherein saidnon-nucleic acid portion has a binding affinity for analytes ofinterest, and wherein when said non-nucleic acid portion is a peptide orprotein, said nucleic acid portion does not comprises sequences whichare either identical or complementary to sequences that code for saidpeptide or protein.
 850. The composition of claim 849, wherein saidsolid surfaces are porous or non-porous.
 851. The composition of claim850, wherein said porous solid surfaces are selected from the groupconsisting of polyacrylamide and agarose.
 852. The composition of claim850, wherein said non-porous solid surfaces comprises glass or plastic.853. The composition of claim 849, wherein said solid surfaces aretransparent, translucent, opaque or reflective.
 854. The composition ofclaim 849, wherein said fixed or immobilized nucleic acid is selectedfrom the group consisting of DNA, RNA and analogs thereof.
 855. Thecomposition of claim 854, wherein said analogs comprise PNA.
 856. Thecomposition of claims 854 or 855, wherein said nucleic acids or analogsare modified on any one of the sugar, phosphate or base moieties. 857.The composition of claim 849, wherein said nucleic acid portion isselected from the group consisting of DNA, RNA and analogs thereof. 858.The composition of claim 857, wherein said analogs comprise PNA. 859.The composition of claims 857 or 858, wherein said nucleic acids oranalogs are modified on any one of the sugar, phosphate or basemoieties.
 860. The composition of claim 849, wherein said non-nucleicacid portions are selected from the group consisting of peptides,proteins, ligands, enzyme substrates, hormones, receptors, drugs and acombination of any of the foregoing.
 861. A process for detecting orquantifying analytes of interest, said process comprising the stepsof: 1) providing: a) an array of solid surfaces comprising a pluralityof discrete areas; wherein at least two of said discrete areas comprisea chimeric composition comprising a nucleic acid portion; and anon-nucleic acid portion; wherein said nucleic acid portion of a firstdiscrete area has the same sequence as the nucleic acid portion of asecond discrete area; and wherein said non-nucleic acid portion has abinding affinity for analytes of interest; b) a sample containing orsuspected of containing one or more of said analytes of interest; and c)signal generating means; 2) contacting said array a) with the sample b)under conditions permissive of binding said analytes to said non-nucleicacid portion; 3) contacting said bound analytes with said signalgenerating means; and 4) detecting or quantifying the presence of saidanalytes.
 862. The process of claim 861, wherein said solid surfaces areporous or non-porous.
 863. The process of claim 862, wherein said poroussolid surfaces are selected from the group consisting of polyacrylamideand agarose.
 864. The process of claim 862, wherein said non-poroussolid surfaces comprise glass or plastic.
 865. The process of claim 861,wherein said solid surfaces are transparent, translucent, opaque orreflective.
 866. The process of claim 861, wherein said nucleic acidportion is selected from the group consisting of DNA, RNA and analogsthereof.
 867. The process of claim 866, wherein said analogs comprisePNA.
 868. The process of claims 866 or 867, wherein said nucleic acidsor analogs are modified on any one of the sugar, phosphate or basemoieties.
 869. The process of claim 861, wherein said nucleic acidportions are directly or indirectly fixed or immobilized to said solidsurfaces.
 870. The process of claim 861, wherein said non-nucleic acidportions are selected from the group consisting of peptides, proteins,ligands, enzyme substrates, hormones, receptors, drugs and a combinationof any of the foregoing.
 871. The process of claim 861, wherein saidsignal generating means comprise direct signal generating means andindirect signal generating means.
 872. The process of claim 871, whereinsaid direct signal generation is selected from the group consisting of afluorescent compound, a phosphorescent compound, a chemiluminescentcompound, a chelating compound, an electron dense compound, a magneticcompound, an intercalating compound, an energy transfer compound and acombination of any of the foregoing.
 873. The process of claim 871,wherein said indirect signal generation is selected from the groupconsisting of an antibody, an antigen, a hapten, a receptor, a hormone,a ligand, an enzyme and a combination of any of the foregoing.
 874. Theprocess of claim 873, wherein said enzyme catalyzes a reaction selectedfrom the group consisting of a fluorogenic reaction, a chromogenicreaction and a chemiluminescent reaction.
 875. A process for detectingor quantifying analytes of interest, said process comprising the stepsof: 1) providing: a) an array of solid surfaces comprising a pluralityof discrete areas; wherein at least two of said discrete areas comprisea chimeric composition comprising a nucleic acid portion; and anon-nucleic acid portion; wherein said nucleic acid portion of a firstdiscrete area has the same sequence as the nucleic acid portion of asecond discrete area; and wherein said non-nucleic acid portion has abinding affinity for analytes of interest; b) a sample containing orsuspected of containing one or more of said analytes of interest; and c)signal generating means; 2) labeling said analytes of interest with saidsignal generating means; 3) contacting said array a) with said labeledanalytes under conditions permissive of binding said labeled analytes tosaid non-nucleic acid portion; and 4) detecting or quantifying thepresence of said analytes.
 876. The process of claim 875, wherein saidsolid surfaces are porous or non-porous.
 877. The process of claim 876,wherein said porous solid surfaces are selected from the groupconsisting of polyacrylamide and agarose.
 878. The process of claim 876,wherein said non-porous solid surfaces comprise glass or plastic. 879.The process of claim 876, wherein said solid surfaces are transparent,translucent, opaque or reflective.
 880. The process of claim 875,wherein said nucleic acid portion is selected from the group consistingof DNA, RNA and analogs thereof.
 881. The process of claim 880, whereinsaid analogs comprise PNA.
 882. The process of claims 880 or 881,wherein said nucleic acids or analogs are modified on any one of thesugar, phosphate or base moieties.
 883. The process of claim 875,wherein said nucleic acid portions are directly or indirectly fixed orimmobilized to said solid surfaces.
 884. The process of claim 875,wherein said non-nucleic acid portions are selected from the groupconsisting of peptides, proteins, ligands, enzyme substrates, hormones,receptors, drugs and a combination of any of the foregoing.
 885. Theprocess of claim 875, wherein said signal generating means comprisedirect signal generating means and indirect signal generating means.886. The process of claim 885, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 887. The process of claim 885, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 888. The process ofclaim 887, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 889. A process for detecting or quantifyinganalytes of interest, said process comprising the steps of: 1) providinga) an array of solid surfaces comprising a plurality of discrete areas;wherein at least two of said discrete areas comprise nucleic acids fixedor immobilized to said discrete areas, b) chimeric compositionscomprising: i) a nucleic acid portion; and ii) a non-nucleic acidportion; said nucleic acid portion comprising at least one sequence,wherein said non-nucleic acid portion has a binding affinity foranalytes of interest, and wherein when said non-nucleic acid portion isa peptide or protein, said nucleic acid portion does not comprisesequences which are either identical or complementary to sequences thatcode for said peptide or protein; c) a sample containing or suspected ofcontaining said analytes of interest; and d) signal generating means; 2)contacting said array with said chimeric compositions to hybridize thenucleic acid portions of said chimeric compositions to complementarynucleic acids fixed or immobilized to said array; 3) contacting saidarray a) with the sample b) under conditions permissive of binding saidanalytes to said non-nucleic acid portion; 4) contacting said boundanalytes with said signal generating means; and 5) detecting orquantifying the presence of said analytes.
 890. The process of claim889, wherein said solid surfaces are porous or non-porous.
 891. Theprocess of claim 890, wherein said porous solid surfaces are selectedfrom the group consisting of polyacrylamide and agarose.
 892. Theprocess of claim 890, wherein said non-porous solid surfaces comprisesglass or plastic.
 893. The process of claim 889, wherein said solidsurfaces are transparent, translucent, opaque or reflective.
 894. Theprocess of claim 889, wherein said fixed or immobilized nucleic acid isselected from the group consisting of DNA, RNA and analogs thereof. 895.The process of claim 894, wherein said analogs comprise PNA.
 896. Theprocess of claims 894 or 895, wherein said nucleic acids or analogs aremodified on any one of the sugar, phosphate or base moieties.
 897. Theprocess of claim 889, wherein said nucleic acid portion is selected fromthe group consisting of DNA, RNA and analogs thereof.
 898. The processof claim 897, wherein said analogs comprise PNA.
 899. The process ofclaims 897 or 898, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 900. The process ofclaim 889, wherein said non-nucleic acid portions are selected from thegroup consisting of peptides, proteins, ligands, enzyme substrates,hormones, receptors, drugs and a combination of any of the foregoing.901. The process of claim 889, wherein said signal generating meanscomprise direct signal generating means and indirect signal generatingmeans.
 902. The process of claim 901, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 903. The process of claim 901, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 904. The process ofclaim 903, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 905. A process for detecting or quantifyinganalytes of interest, said process comprising the steps of: 1) providinga) an array of solid surfaces comprising a plurality of discrete areas;wherein at least two of said discrete areas comprise nucleic acids fixedor immobilized to said discrete areas, b) chimeric compositionscomprising: i) a nucleic acid portion; and ii) a non-nucleic acidportion; said nucleic acid portion comprising at least one sequence,wherein said non-nucleic acid portion has a binding affinity foranalytes of interest, and wherein when said non-nucleic acid portion isa peptide or protein, said nucleic acid portion does not comprisesequences which are either identical or complementary to sequences thatcode for said peptide or protein; c) a sample containing or suspected ofcontaining said analytes of interest; and d) signal generating means; 2)contacting said chimeric compositions with the sample b) underconditions permissive of binding said analytes to said non-nucleic acidportion; 3) contacting said array with said chimeric compositions tohybridize the nucleic acid portions of said chimeric compositions tocomplementary nucleic acids fixed or immobilized to said array; 4)contacting said bound analytes with said signal generating means; and 5)detecting or quantifying the presence of said analytes.
 906. The processof claim 905, wherein said solid surfaces are porous or non-porous. 907.The process of claim 906, wherein said porous solid surfaces areselected from the group consisting of polyacrylamide and agarose. 908.The process of claim 906, wherein said non-porous solid surfacescomprises glass or plastic.
 909. The process of claim 905, wherein saidsolid surfaces are transparent, translucent, opaque or reflective. 910.The process of claim 905, wherein said fixed or immobilized nucleic acidis selected from the group consisting of DNA, RNA and analogs thereof.911. The process of claim 910, wherein said analogs comprise PNA. 912.The process of claims 910 or 911, wherein said nucleic acids or analogsare modified on any one of the sugar, phosphate or base moieties. 913.The process of claim 905, wherein said nucleic acid portion is selectedfrom the group consisting of DNA, RNA and analogs thereof.
 914. Theprocess of claim 913, wherein said analogs comprise PNA.
 915. Theprocess of claims 913 or 914, wherein said nucleic acids or analogs aremodified on any one of the sugar, phosphate or base moieties.
 916. Theprocess of claim 905, wherein said non-nucleic acid portions areselected from the group consisting of peptides, proteins, ligands,enzyme substrates, hormones, receptors, drugs and a combination of anyof the foregoing.
 917. The process of claim 905, wherein said signalgenerating means comprise direct signal generating means and indirectsignal generating means.
 918. The process of claim 917, wherein saiddirect signal generation is selected from the group consisting of afluorescent compound, a phosphorescent compound, a chemiluminescentcompound, a chelating compound, an electron dense compound, a magneticcompound, an intercalating compound, an energy transfer compound and acombination of any of the foregoing.
 919. The process of claim 917,wherein said indirect signal generation is selected from the groupconsisting of an antibody, an antigen, a hapten, a receptor, a hormone,a ligand, an enzyme and a combination of any of the foregoing.
 920. Theprocess of claim 919, wherein said enzyme catalyzes a reaction selectedfrom the group consisting of a fluorogenic reaction, a chromogenicreaction and a chemiluminescent reaction.
 921. A process for detectingor quantifying analytes of interest, said process comprising the stepsof: 1) providing a) an array of solid surfaces comprising a plurality ofdiscrete areas; wherein at least two of said discrete areas comprisenucleic acids fixed or immobilized to said discrete areas, b) chimericcompositions comprising: i) a nucleic acid portion; and ii) anon-nucleic acid portion; said nucleic acid portion comprising at leastone sequence, wherein said non-nucleic acid portion has a bindingaffinity for analytes of interest, and wherein when said non-nucleicacid portion is a peptide or protein, said nucleic acid portion does notcomprises sequences which are either identical or complementary tosequences that code for said peptide or protein; c) a sample containingor suspected of containing said analytes of interest; and d) signalgenerating means; 2) contacting said array with said chimericcompositions to hybridize the nucleic acid portions of said chimericcompositions to complementary nucleic acids fixed or immobilized to saidarray; 3) labeling said analytes of interest with said signal generatingmeans; 4) contacting said array with the labeled analytes to bind saidanalytes to said non-nucleic acid portion; and 5) detecting orquantifying the presence of said analytes.
 922. The process of claim921, wherein said solid surfaces are porous or non-porous.
 923. Theprocess of claim 922, wherein said porous solid surfaces are selectedfrom the group consisting of polyacrylamide and agarose.
 924. Theprocess of claim 922, wherein said non-porous solid surfaces comprisesglass or plastic.
 925. The process of claim 921, wherein said solidsurfaces are transparent, translucent, opaque or reflective.
 926. Theprocess of claim 921, wherein said fixed or immobilized nucleic acid isselected from the group consisting of DNA, RNA and analogs thereof. 927.The process of claim 926, wherein said analogs comprise PNA.
 928. Theprocess of claims 926 or 927, wherein said nucleic acids or analogs aremodified on any one of the sugar, phosphate or base moieties.
 929. Theprocess of claim 921, wherein said nucleic acid portion is selected fromthe group consisting of DNA, RNA and analogs thereof.
 930. The processof claim 929, wherein said analogs comprise PNA.
 931. The process ofclaims 929 or 930, wherein said nucleic acids or analogs are modified onany one of the sugar, phosphate or base moieties.
 932. The process ofclaim 921, wherein said non-nucleic acid portions are selected from thegroup consisting of peptides, proteins, ligands, enzyme substrates,hormones, receptors, drugs and a combination of any of the foregoing.933. The process of claim 921, wherein said signal generating meanscomprise direct signal generating means and indirect signal generatingmeans.
 934. The process of claim 933, wherein said direct signalgeneration is selected from the group consisting of a fluorescentcompound, a phosphorescent compound, a chemiluminescent compound, achelating compound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 935. The process of claim 933, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 936. The process ofclaim 935, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.
 937. A process for detecting or quantifyinganalytes of interest, said process comprising the steps of: 1) providinga) an array of solid surfaces comprising a plurality of discrete areas;wherein at least two of said discrete areas comprise nucleic acids fixedor immobilized to said discrete areas, b) chimeric compositionscomprising: i) a nucleic acid portion; and ii) a non-nucleic acidportion; said nucleic acid portion comprising at least one sequence,wherein said non-nucleic acid portion has a binding affinity foranalytes of interest, and wherein when said non-nucleic acid portion isa peptide or protein, said nucleic acid portion does not comprisessequences which are either identical or complementary to sequences thatcode for said peptide or protein; c) a sample containing or suspected ofcontaining said analytes of interest; and d) signal generating means; 2)labeling said analytes of interest with said signal generating means; 3)contacting said chimeric compositions with the labeled analytes to bindsaid analytes to said non-nucleic acid portion; 4) contacting said arraywith said chimeric compositions to hybridize the nucleic acid portionsof said chimeric compositions to complementary nucleic acids fixed orimmobilized to said array; and 5) detecting or quantifying the presenceof said analytes.
 938. The process of claim 937, wherein said solidsurfaces are porous or non-porous.
 939. The process of claim 938,wherein said porous solid surfaces are selected from the groupconsisting of polyacrylamide and agarose.
 940. The process of claim 938,wherein said non-porous solid surfaces comprises glass or plastic. 941.The process of claim 937, wherein said solid surfaces are transparent,translucent, opaque or reflective.
 942. The process of claim 937,wherein said fixed or immobilized nucleic acid is selected from thegroup consisting of DNA, RNA and analogs thereof.
 943. The process ofclaim 942, wherein said analogs comprise PNA.
 944. The process of claims942 or 943, wherein said nucleic acids or analogs are modified on anyone of the sugar, phosphate or base moieties.
 945. The process of claim937, wherein said nucleic acid portion is selected from the groupconsisting of DNA, RNA and analogs thereof.
 946. The process of claim945, wherein said analogs comprise PNA.
 947. The process of claims 945or 946, wherein said nucleic acids or analogs are modified on any one ofthe sugar, phosphate or base moieties.
 948. The process of claim 937,wherein said non-nucleic acid portions are selected from the groupconsisting of peptides, proteins, ligands, enzyme substrates, hormones,receptors, drugs and a combination of any of the foregoing.
 949. Theprocess of claim 937, wherein said signal generating means comprisedirect signal generating means and indirect signal generating means.950. The process of claim 949, wherein said direct signal generation isselected from the group consisting of a fluorescent compound, aphosphorescent compound, a chemiluminescent compound, a chelatingcompound, an electron dense compound, a magnetic compound, anintercalating compound, an energy transfer compound and a combination ofany of the foregoing.
 951. The process of claim 949, wherein saidindirect signal generation is selected from the group consisting of anantibody, an antigen, a hapten, a receptor, a hormone, a ligand, anenzyme and a combination of any of the foregoing.
 952. The process ofclaim 951, wherein said enzyme catalyzes a reaction selected from thegroup consisting of a fluorogenic reaction, a chromogenic reaction and achemiluminescent reaction.